Modulators of atp-binding cassette transporters

ABSTRACT

Compounds of the present invention and pharmaceutically acceptable compositions thereof, are useful as modulators of ATP-Binding Cassette (“ABC”) transporters or fragments thereof, including Cystic Fibrosis Transmembrane Conductance Regulator (“CFTR”). The present invention also relates to methods of treating ABC transporter mediated diseases using compounds of the present invention.

CLAIM OF PRIORITY

The present patent application claims priority to U.S. provisionalpatent application Ser. No. 60/790,459, filed on Apr. 7, 2006, which ishereby incorporated by reference in its entirety.

TECHNICAL FIELD OF THE INVENTION

The present invention relates to modulators of ATP-Binding Cassette(“ABC”) transporters or fragments thereof, including Cystic FibrosisTransmembrane Conductance Regulator (“CFTR”), compositions thereof andmethods therewith. The present invention also relates to methods oftreating ABC transporter mediated diseases using such modulators.

BACKGROUND OF THE INVENTION

ABC transporters are a family of membrane transporter proteins thatregulate the transport of a wide variety of pharmacological agents,potentially toxic drugs, and xenobiotics, as well as anions. ABCtransporters are homologous membrane proteins that bind and use cellularadenosine triphosphate (ATP) for their specific activities. Some ofthese transporters were discovered as multidrug resistance proteins(like the MDR1-P glycoprotein, or the multidrug resistance protein,MRP1), defending malignant cancer cells against chemotherapeutic agents.To date, 48 ABC Transporters have been identified and grouped into 7families based on their sequence identity and function.

ABC transporters regulate a variety of important physiological roleswithin the body and provide defense against harmful environmentalcompounds. Because of this, they represent important potential drugtargets for the treatment of diseases associated with defects in thetransporter, prevention of drug transport out of the target cell, andintervention in other diseases in which modulation of ABC transporteractivity may be beneficial.

One member of the ABC transporter family commonly associated withdisease is the cAMP/ATP-mediated anion channel, CFTR. CFTR is expressedin a variety of cells types, including absorptive and secretoryepithelia cells, where it regulates anion flux across the membrane, aswell as the activity of other ion channels and proteins. In epitheliacells, normal functioning of CFTR is critical for the maintenance ofelectrolyte transport throughout the body, including respiratory anddigestive tissue. CFTR is composed of approximately 1480 amino acidsthat encode a protein made up of a tandem repeat of transmembranedomains, each containing six transmembrane helices and a nucleotidebinding domain. The two transmembrane domains are linked by a large,polar, regulatory (R)-domain with multiple phosphorylation sites thatregulate channel activity and cellular trafficking.

The gene encoding CFTR has been identified and sequenced (See Gregory,R. J. et al. (1990) Nature 347:382-386; Rich, D. P. et al. (1990) Nature347:358-362), (Riordan, J. R. et al. (1989) Science 245:1066-1073). Adefect in this gene causes mutations in CFTR resulting in CysticFibrosis (“CF”), the most common fatal genetic disease in humans. CysticFibrosis affects approximately one in every 2,500 infants in the UnitedStates. Within the general United States population, up to 10 millionpeople carry a single copy of the defective gene without apparent illeffects. In contrast, individuals with two copies of the CF associatedgene suffer from the debilitating and fatal effects of CF, includingchronic lung disease.

In patients with cystic fibrosis, mutations in CFTR endogenouslyexpressed in respiratory epithelia leads to reduced apical anionsecretion causing an imbalance in ion and fluid transport. The resultingdecrease in anion transport contributes to enhanced mucus accumulationin the lung and the accompanying microbial infections that ultimatelycause death in CF patients. In addition to respiratory disease, CFpatients typically suffer from gastrointestinal problems and pancreaticinsufficiency that, if left untreated, results in death. In addition,the majority of males with cystic fibrosis are infertile and fertilityis decreased among females with cystic fibrosis. In contrast to thesevere effects of two copies of the CF associated gene, individuals witha single copy of the CF associated gene exhibit increased resistance tocholera and to dehydration resulting from diarrhea—perhaps explainingthe relatively high frequency of the CF gene within the population.

Sequence analysis of the CFTR gene of CF chromosomes has revealed avariety of disease causing mutations (Cutting, G. R. et al. (1990)Nature 346:366-369; Dean, M. et al. (1990) Cell 61:863:870; and Kerem,B-S. et al. (1989) Science 245:1073-1080; Kerem, B-S et al. (1990) Proc.Natl. Acad. Sci. USA 87:8447-8451). To date, >1000 disease causingmutations in the CF gene have been identified(http://www.genet.sickkids.on.ca/cftr/). The most prevalent mutation isa deletion of phenylalanine at position 508 of the CFTR amino acidsequence, and is commonly referred to as ΔF508-CFTR. This mutationoccurs in approximately 70% of the cases of cystic fibrosis and isassociated with a severe disease.

The deletion of residue 508 in ΔF508-CFTR prevents the nascent proteinfrom folding correctly. This results in the inability of the mutantprotein to exit the ER, and traffic to the plasma membrane. As a result,the number of channels present in the membrane is far less than observedin cells expressing wild-type CFTR. In addition to impaired trafficking,the mutation results in defective channel gating. Together, the reducednumber of channels in the membrane and the defective gating lead toreduced anion transport across epithelia leading to defective ion andfluid transport. (Quinton, P. M. (1990), FASEB J. 4: 2709-2727). Studieshave shown, however, that the reduced numbers of ΔF508-CFTR in themembrane are functional, albeit less than wild-type CFTR. (Dalemans etal. (1991), Nature Lond. 354: 526-528; Denning et al., supra; Pasyk andFoskett (1995), J. Cell. Biochem. 270: 12347-50). In addition toΔF508-CFTR, other disease causing mutations in CFTR that result indefective trafficking, synthesis, and/or channel gating could be up- ordown-regulated to alter anion secretion and modify disease progressionand/or severity.

Although CFTR transports a variety of molecules in addition to anions,it is clear that this role (the transport of anions) represents oneelement in an important mechanism of transporting ions and water acrossthe epithelium. The other elements include the epithelial Na⁺ channel,ENaC, Na⁺/2Cl⁻/K⁺ co-transporter, Na⁺—K⁺-ATPase pump and the basolateralmembrane K⁺ channels, that are responsible for the uptake of chlorideinto the cell.

These elements work together to achieve directional transport across theepithelium via their selective expression and localization within thecell. Chloride absorption takes place by the coordinated activity ofENaC and CFTR present on the apical membrane and the Na⁺—K⁺-ATPase pumpand Cl− channels expressed on the basolateral surface of the cell.Secondary active transport of chloride from the luminal side leads tothe accumulation of intracellular chloride, which can then passivelyleave the cell via Cl⁻ channels, resulting in a vectorial transport.Arrangement of Na⁺/2Cl⁻/K⁺ co-transporter, Na⁺—K⁺-ATPase pump and thebasolateral membrane K⁺ channels on the basolateral surface and CFTR onthe luminal side coordinate the secretion of chloride via CFTR on theluminal side. Because water is probably never actively transporteditself, its flow across epithelia depends on tiny transepithelialosmotic gradients generated by the bulk flow of sodium and chloride.

In addition to Cystic Fibrosis, modulation of CFTR activity may bebeneficial for other diseases not directly caused by mutations in CFTR,such as secretory diseases and other protein folding diseases mediatedby CFTR. These include, but are not limited to, chronic obstructivepulmonary disease (COPD), dry eye disease, and Sjögren's Syndrome.

COPD is characterized by airflow limitation that is progressive and notfully reversible. The airflow limitation is due to mucus hypersecretion,emphysema, and bronchiolitis. Activators of mutant or wild-type CFTRoffer a potential treatment of mucus hypersecretion and impairedmucociliary clearance that is common in COPD. Specifically, increasinganion secretion across CFTR may facilitate fluid transport into theairway surface liquid to hydrate the mucus and optimized periciliaryfluid viscosity. This would lead to enhanced mucociliary clearance and areduction in the symptoms associated with COPD. Dry eye disease ischaracterized by a decrease in tear aqueous production and abnormal tearfilm lipid, protein and mucin profiles. There are many causes of dryeye, some of which include age, Lasik eye surgery, arthritis,medications, chemical/thermal burns, allergies, and diseases, such asCystic Fibrosis and Sjögrens's syndrome. Increasing anion secretion viaCFTR would enhance fluid transport from the corneal endothelial cellsand secretory glands surrounding the eye to increase corneal hydration.This would help to alleviate the symptoms associated with dry eyedisease. Sjögrens's syndrome is an autoimmune disease in which theimmune system attacks moisture-producing glands throughout the body,including the eye, mouth, skin, respiratory tissue, liver, vagina, andgut. Symptoms, include, dry eye, mouth, and vagina, as well as lungdisease. The disease is also associated with rheumatoid arthritis,systemic lupus, systemic sclerosis, and polymypositis/dermatomyositis.Defective protein trafficking is believed to cause the disease, forwhich treatment options are limited. Modulators of CFTR activity mayhydrate the various organs afflicted by the disease and help to elevatethe associated symptoms.

As discussed above, it is believed that the deletion of residue 508 inΔF508-CFTR prevents the nascent protein from folding correctly,resulting in the inability of this mutant protein to exit the ER, andtraffic to the plasma membrane. As a result, insufficient amounts of themature protein are present at the plasma membrane and chloride transportwithin epithelial tissues is significantly reduced. In fact, thiscellular phenomenon of defective ER processing of ABC transporters bythe ER machinery has been shown to be the underlying basis not only forCF disease, but for a wide range of other isolated and inheriteddiseases. The two ways that the ER machinery can malfunction is eitherby loss of coupling to ER export of the proteins leading to degradation,or by the ER accumulation of these defective/misfolded proteins [AridorM, et al., Nature Med., 5(7), pp 745-751 (1999); Shastry, B. S., et al.,Neurochem. International, 43, pp 1-7 (2003); Rutishauser, J., et al.,Swiss Med Wkly, 132, pp 211-222 (2002); Morello, J P et al., TIPS, 21,pp. 466-469 (2000); Bross P., et al., Human Mut., 14, pp. 186-198(1999)]. The diseases associated with the first class of ER malfunctionare Cystic fibrosis (due to misfolded ΔF508-CFTR as discussed above),Hereditary emphysema (due to al-antitrypsin; non Piz variants),Hereditary hemochromatosis, Coagulation-Fibrinolysis deficiencies, suchas Protein C deficiency, Type 1 hereditary angioedema, Lipid processingdeficiencies, such as Familial hypercholesterolemia, Type 1chylomicronemia, Abetalipoproteinemia, Lysosomal storage diseases, suchas I-cell disease/Pseudo-Hurler, Mucopolysaccharidoses (due to Lysosomalprocessing enzymes), Sandhof/Tay-Sachs (due to β-Hexosaminidase),Crigler-Najjar type II (due to UDP-glucuronyl-sialyc-transferase),Polyendocrinopathy/Hyperinsulemia, Diabetes mellitus (due to Insulinreceptor), Laron dwarfism (due to Growth hormone receptor),Myleoperoxidase deficiency, Primary hypoparathyroidism (due toPreproparathyroid hormone), Melanoma (due to Tyrosinase). The diseasesassociated with the latter class of ER malfunction are Glycanosis CDGtype 1, Hereditary emphysema (due to al-Antitrypsin (PiZ variant),Congenital hyperthyroidism, Osteogenesis imperfecta (due to Type I, II,IV procollagen), Hereditary hypofibrinogenemia (due to Fibrinogen), ACTdeficiency (due to al-Antichymotrypsin), Diabetes insipidus (DI),Neurophyseal DI (due to Vasopvessin hormone/V2-receptor), Neprogenic DI(due to Aquaporin II), Charcot-Marie Tooth syndrome (due to Peripheralmyelin protein 22), Perlizaeus-Merzbacher disease, neurodegenerativediseases such as Alzheimer's disease (due to PAPP and presenilins),Parkinson's disease, Amyotrophic lateral sclerosis, Progressivesupranuclear plasy, Pick's disease, several polyglutamine neurologicaldisorders asuch as Huntington, Spinocerebullar ataxia type I, Spinal andbulbar muscular atrophy, Dentatorubal pallidoluysian, and Myotonicdystrophy, as well as Spongiform encephalopathies, such as HereditaryCreutzfeldt-Jakob disease (due to Prion protein processing defect),Fabry disease (due to lysosomal a-galactosidase A) andStraussler-Scheinker syndrome (due to Prp processing defect).

In addition to up-regulation of CFTR activity, reducing anion secretionby CFTR modulators may be beneficial for the treatment of secretorydiarrheas, in which epithelial water transport is dramatically increasedas a result of secretagogue activated chloride transport. The mechanisminvolves elevation of cAMP and stimulation of CFTR.

Although there are numerous causes of diarrhea, the major consequencesof diarrheal diseases, resulting from excessive chloride transport arecommon to all, and include dehydration, acidosis, impaired growth anddeath.

Acute and chronic diarrheas represent a major medical problem in manyareas of the world. Diarrhea is both a significant factor inmalnutrition and the leading cause of death (5,000,000 deaths/year) inchildren less than five years old.

Secretory diarrheas are also a dangerous condition in patients ofacquired immunodeficiency syndrome (AIDS) and chronic inflammatory boweldisease (IBD). 16 million travelers to developing countries fromindustrialized nations every year develop diarrhea, with the severityand number of cases of diarrhea varying depending on the country andarea of travel.

Diarrhea in bam animals and pets such as cows, pigs and horses, sheep,goats, cats and dogs, also known as scours, is a major cause of death inthese animals. Diarrhea can result from any major transition, such asweaning or physical movement, as well as in response to a variety ofbacterial or viral infections and generally occurs within the first fewhours of the animal's life.

The most common diarrhea causing bacteria is enterotoxogenic E. coli(ETEC) having the K99 pilus antigen. Common viral causes of diarrheainclude rotavirus and coronavirus. Other infectious agents includecryptosporidium, giardia lamblia, and salmonella, among others.

Symptoms of rotaviral infection include excretion of watery feces,dehydration and weakness. Coronavirus causes a more severe illness inthe newborn animals, and has a higher mortality rate than rotaviralinfection. Often, however, a young animal may be infected with more thanone virus or with a combination of viral and bacterial microorganisms atone time. This dramatically increases the severity of the disease.

Accordingly, there is a need for modulators of an ABC transporteractivity, and compositions thereof, that can be used to modulate theactivity of the ABC transporter in the cell membrane of a mammal.

There is a need for methods of treating ABC transporter mediateddiseases using such modulators of ABC transporter activity.

There is a need for methods of modulating an ABC transporter activity inan ex vivo cell membrane of a mammal.

There is a need for modulators of CFTR activity that can be used tomodulate the activity of CFTR in the cell membrane of a mammal.

There is a need for methods of treating CFTR-mediated diseases usingsuch modulators of CFTR activity.

There is a need for methods of modulating CFTR activity in an ex vivocell membrane of a mammal.

SUMMARY OF THE INVENTION

It has now been found that compounds of this invention, andpharmaceutically acceptable compositions thereof, are useful asmodulators of ABC transporter activity, particularly CTFR activity.These compounds have the general formula I:

or a pharmaceutically acceptable salt thereof, wherein R₁, R₂, ring A,ring B, and n are defined below.

These compounds and pharmaceutically acceptable compositions are usefulfor treating or lessening the severity of a variety of diseases,disorders, or conditions, including, but not limited to, cysticfibrosis, hereditary emphysema, hereditary hemochromatosis,coagulation-fibrinolysis deficiencies, such as protein C deficiency,Type 1 hereditary angioedema, lipid processing deficiencies, such asfamilial hypercholesterolemia, Type 1 chylomicronemia,abetalipoproteinemia, lysosomal storage diseases, such as I-celldisease/pseudo-Hurler, mucopolysaccharidoses, Sandhof/Tay-Sachs,Crigler-Najjar type II, polyendocrinopathy/hyperinsulemia, diabetesmellitus, laron dwarfism, myleoperoxidase deficiency, primaryhypoparathyroidism, melanoma, glycanosis CDG type 1, hereditaryemphysema, congenital hyperthyroidism, osteogenesis imperfecta,hereditary hypofibrinogenemia, ACT deficiency, diabetes insipidus,neurophysiol, nephrogenic, Charcot-Marie Tooth syndrome,Perlizaeus-Merzbacher disease, neurodegenerative diseases such asAlzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis,progressive supranuclear plasy, Pick's disease, several polyglutamineneurological disorders asuch as Huntington, spinocerebullar ataxia typeI, spinal and bulbar muscular atrophy, dentatorubal pallidoluysian, andmyotonic dystrophy, as well as spongiform encephalopathies, such ashereditary Creutzfeldt-Jakob disease, Fabry disease,Straussler-Scheinker syndrome, COPD, dry-eye disease, and Sjögren'sdisease.

DETAILED DESCRIPTION OF THE INVENTION I. Definitions

As used herein, the following definitions shall apply unless otherwiseindicated.

The term “ABC-transporter” as used herein means an ABC-transporterprotein or a fragment thereof comprising at least one binding domain,wherein said protein or fragment thereof is present in vivo or in vitro.The term “binding domain” as used herein means a domain on theABC-transporter that can bind to a modulator. See, e.g., Hwang, T. C. etal., J. Gen. Physiol. (1998): 111(3), 477-90.

The term “CFTR” as used herein means cystic fibrosis transmembraneconductance regulator or a mutation thereof capable of regulatoractivity, including, but not limited to, ΔF508 CFTR and G551D CFTR (see,e.g., http://www.genet.sickkids.on.ca/cftr/, for CFTR mutations).

The term “modulating” as used herein means increasing or decreasing,e.g. activity, by a measurable amount. Compounds that modulate ABCTransporter activity, such as CFTR activity, by increasing the activityof the ABC Transporter, e.g., a CFTR anion channel, are called agonists.Compounds that modulate ABC Transporter activity, such as CFTR activity,by decreasing the activity of the ABC Transporter, e.g., CFTR anionchannel, are called antagonists. An agonist interacts with an ABCTransporter, such as CFTR anion channel, to increase the ability of thereceptor to transduce an intracellular signal in response to endogenousligand binding. An antagonist interacts with an ABC Transporter, such asCFTR, and competes with the endogenous ligand(s) or substrate(s) forbinding site(s) on the receptor to decrease the ability of the receptorto transduce an intracellular signal in response to endogenous ligandbinding.

The phrase “treating or reducing the severity of an ABC Transportermediated disease” refers both to treatments for diseases that aredirectly caused by ABC Transporter and/or CFTR activities andalleviation of symptoms of diseases not directly caused by ABCTransporter and/or CFTR anion channel activities. Examples of diseaseswhose symptoms may be affected by ABC Transporter and/or CFTR activityinclude, but are not limited to, Cystic fibrosis, Hereditary emphysema,Hereditary hemochromatosis, Coagulation-Fibrinolysis deficiencies, suchas Protein C deficiency, Type 1 hereditary angioedema, Lipid processingdeficiencies, such as Familial hypercholesterolemia, Type 1chylomicronemia, Abetalipoproteinemia, Lysosomal storage diseases, suchas I-cell disease/Pseudo-Hurler, Mucopolysaccharidoses,Sandhof/Tay-Sachs, Crigler-Najjar type II,Polyendocrinopathy/Hyperinsulemia, Diabetes mellitus, Laron dwarfism,Myleoperoxidase deficiency, Primary hypoparathyroidism, Melanoma,Glycanosis CDG type 1, Hereditary emphysema, Congenital hyperthyroidism,Osteogenesis imperfecta, Hereditary hypofibrinogenemia, ACT deficiency,Diabetes insipidus (DI), Neurophysiol DI, Nephrogenic DI, Charcot-MarieTooth syndrome, Perlizaeus-Merzbacher disease, neurodegenerativediseases such as Alzheimer's disease, Parkinson's disease, Amyotrophiclateral sclerosis, Progressive supranuclear plasy, Pick's disease,several polyglutamine neurological disorders such as Huntington,Spinocerebullar ataxia type I, Spinal and bulbar muscular atrophy,Dentatorubal pallidoluysian, and Myotonic dystrophy, as well asSpongiform encephalopathies, such as Hereditary Creutzfeldt-Jakobdisease, Fabry disease, Straussler-Scheinker syndrome, COPD, dry-eyedisease, and Sjogren's disease.

For purposes of this invention, the chemical elements are identified inaccordance with the Periodic Table of the Elements, CAS version,Handbook of Chemistry and Physics, 75th Ed. Additionally, generalprinciples of organic chemistry are described in “Organic Chemistry”,Thomas Sorrell, University Science Books, Sausolito: 1999, and “March'sAdvanced Organic Chemistry”, 5th Ed., Ed.: Smith, M. B. and March, J.,John Wiley & Sons, New York: 2001, the entire contents of which arehereby incorporated by reference.

As described herein, compounds of the invention may optionally besubstituted with one or more substituents, such as are illustratedgenerally above, or as exemplified by particular classes, subclasses,and species of the invention.

As used herein the term “aliphatic” encompasses the terms alkyl,alkenyl, alkynyl, each of which being optionally substituted as setforth below.

As used herein, an “alkyl” group refers to a saturated aliphatichydrocarbon group containing 1-12 (e.g., 1-8, 1-6, or 1-4) carbon atoms.An alkyl group can be straight or branched. Examples of alkyl groupsinclude, but are not limited to, methyl, ethyl, propyl, isopropyl,butyl, isobutyl, sec-butyl, tert-butyl, n-pentyl, n-heptyl, or2-ethylhexyl. An alkyl group can be substituted (i.e., optionallysubstituted) with one or more substituents such as halo, phospho,cycloaliphatic [e.g., cycloalkyl or cycloalkenyl], heterocycloaliphatic[e.g., heterocycloalkyl or heterocycloalkenyl], aryl, heteroaryl,alkoxy, aroyl, heteroaroyl, acyl [e.g., (aliphatic)carbonyl,(cycloaliphatic)carbonyl, or (heterocycloaliphatic)carbonyl], nitro,cyano, amido [e.g., (cycloalkylalkyl)carbonylamino, arylcarbonylamino,aralkylcarbonylamino, (heterocycloalkyl)carbonylamino,(heterocycloalkylalkyl)carbonylamino, heteroarylcarbonylamino,heteroaralkylcarbonylamino alkylaminocarbonyl, cycloalkylaminocarbonyl,heterocycloalkylaminocarbonyl, arylaminocarbonyl, orheteroarylaminocarbonyl], amino [e.g., aliphaticamino,cycloaliphaticamino, or heterocycloaliphaticamino], sulfonyl [e.g.,aliphatic-SO₂-], sulfinyl, sulfanyl, sulfoxy, urea, thiourea, sulfamoyl,sulfamide, oxo, carboxy, carbamoyl, cycloaliphaticoxy,heterocycloaliphaticoxy, aryloxy, heteroaryloxy, aralkyloxy,heteroarylalkoxy, alkoxycarbonyl, alkylcarbonyloxy, or hydroxy. Withoutlimitation, some examples of substituted alkyls include carboxyalkyl(such as HOOC-alkyl, alkoxycarbonylalkyl, and alkylcarbonyloxyalkyl),cyanoalkyl, hydroxyalkyl, alkoxyalkyl, acylalkyl, aralkyl,(alkoxyaryl)alkyl, (sulfonylamino)alkyl (such as(alkyl-SO₂-amino)alkyl), aminoalkyl, amidoalkyl, (cycloaliphatic)alkyl,or haloalkyl.

As used herein, an “alkenyl” group refers to an aliphatic carbon groupthat contains 2-8 (e.g., 2-12, 2-6, or 2-4) carbon atoms and at leastone double bond. Like an alkyl group, an alkenyl group can be straightor branched. Examples of an alkenyl group include, but are not limitedto allyl, isoprenyl, 2-butenyl, and 2-hexenyl. An alkenyl group can beoptionally substituted with one or more substituents such as halo,phospho, cycloaliphatic [e.g., cycloalkyl or cycloalkenyl],heterocycloaliphatic [e.g., heterocycloalkyl or heterocycloalkenyl],aryl, heteroaryl, alkoxy, aroyl, heteroaroyl, acyl [e.g.,(aliphatic)carbonyl, (cycloaliphatic)carbonyl, or(heterocycloaliphatic)carbonyl], nitro, cyano, amido [e.g.,(cycloalkylalkyl)carbonylamino, arylcarbonylamino, aralkylcarbonylamino,(heterocycloalkyl)carbonylamino, (heterocycloalkylalkyl)carbonylamino,heteroarylcarbonylamino, heteroaralkylcarbonylamino alkylaminocarbonyl,cycloalkylaminocarbonyl, heterocycloalkylaminocarbonyl,arylaminocarbonyl, or heteroarylaminocarbonyl], amino [e.g.,aliphaticamino, cycloaliphaticamino, heterocycloaliphaticamino, oraliphaticsulfonylamino], sulfonyl [e.g., alkyl-SO₂—,cycloaliphatic-SO₂—, or aryl-SO₂-], sulfinyl, sulfanyl, sulfoxy, urea,thiourea, sulfamoyl, sulfamide, oxo, carboxy, carbamoyl,cycloaliphaticoxy, heterocycloaliphaticoxy, aryloxy, heteroaryloxy,aralkyloxy, heteroaralkoxy, alkoxycarbonyl, alkylcarbonyloxy, orhydroxy. Without limitation, some examples of substituted alkenylsinclude cyanoalkenyl, alkoxyalkenyl, acylalkenyl, hydroxyalkenyl,aralkenyl, (alkoxyaryl)alkenyl, (sulfonylamino)alkenyl (such as(alkyl-SO₂-amino)alkenyl), aminoalkenyl, amidoalkenyl,(cycloaliphatic)alkenyl, or haloalkenyl.

As used herein, an “alkynyl” group refers to an aliphatic carbon groupthat contains 2-8 (e.g., 2-12, 2-6, or 2-4) carbon atoms and has atleast one triple bond. An alkynyl group can be straight or branched.Examples of an alkynyl group include, but are not limited to, propargyland butynyl. An alkynyl group can be optionally substituted with one ormore substituents such as aroyl, heteroaroyl, alkoxy, cycloalkyloxy,heterocycloalkyloxy, aryloxy, heteroaryloxy, aralkyloxy, nitro, carboxy,cyano, halo, hydroxy, sulfo, mercapto, sulfanyl [e.g., aliphaticsulfanylor cycloaliphaticsulfanyl], sulfinyl [e.g., aliphaticsulfinyl orcycloaliphaticsulfinyl], sulfonyl [e.g., aliphatic-SO₂—,aliphaticamino-SO₂—, or cycloaliphatic-SO₂-], amido [e.g.,aminocarbonyl, alkylaminocarbonyl, alkylcarbonylamino,cycloalkylaminocarbonyl, heterocycloalkylaminocarbonyl,cycloalkylcarbonylamino, arylaminocarbonyl, arylcarbonylamino,aralkylcarbonylamino, (heterocycloalkyl)carbonylamino,(cycloalkylalkyl)carbonylamino, heteroaralkylcarbonylamino,heteroarylcarbonylamino or heteroarylaminocarbonyl], urea, thiourea,sulfamoyl, sulfamide, alkoxycarbonyl, alkylcarbonyloxy, cycloaliphatic,heterocycloaliphatic, aryl, heteroaryl, acyl [e.g.,(cycloaliphatic)carbonyl or (heterocycloaliphatic)carbonyl], amino[e.g., aliphaticamino], sulfoxy, oxo, carboxy, carbamoyl,(cycloaliphatic)oxy, (heterocycloaliphatic)oxy, or (heteroaryl)alkoxy.

As used herein, an “amido” encompasses both “aminocarbonyl” and“carbonylamino”. These terms when used alone or in connection withanother group refer to an amido group such as —N(R^(X))—C(O)—R^(Y) or—C(O)—N(R^(X))₂, when used terminally, and —C(O)—N(R^(X))— or—N(R^(X))—C(O)— when used internally, wherein R^(X) and R^(Y) aredefined below. Examples of amido groups include alkylamido (such asalkylcarbonylamino or alkylaminocarbonyl), (heterocycloaliphatic)amido,(heteroaralkyl)amido, (heteroaryl)amido, (heterocycloalkyl)alkylamido,arylamido, aralkylamido, (cycloalkyl)alkylamido, or cycloalkylamido.

As used herein, an “amino” group refers to —NR^(X)R^(Y) wherein each ofR^(X) and R^(Y) is independently hydrogen, aliphatic, cycloaliphatic,(cycloaliphatic)aliphatic, aryl, araliphatic, heterocycloaliphatic,(heterocycloaliphatic)aliphatic, heteroaryl, carboxy, sulfanyl,sulfinyl, sulfonyl, (aliphatic)carbonyl, (cycloaliphatic)carbonyl,((cycloaliphatic)aliphatic)carbonyl, arylcarbonyl,(araliphatic)carbonyl, (heterocycloaliphatic)carbonyl,((heterocycloaliphatic)aliphatic)carbonyl, (heteroaryl)carbonyl, or(heteroaraliphatic)carbonyl, each of which being defined herein andbeing optionally substituted. Examples of amino groups includealkylamino, dialkylamino, or arylamino. When the term “amino” is not theterminal group (e.g., alkylcarbonylamino), it is represented by—NR^(X)—. R^(X) has the same meaning as defined above.

As used herein, an “aryl” group used alone or as part of a larger moietyas in “aralkyl”, “aralkoxy”, or “aryloxyalkyl” refers to monocyclic(e.g., phenyl); bicyclic (e.g., indenyl, naphthalenyl,tetrahydronaphthyl, tetrahydroindenyl); and tricyclic (e.g., fluorenyltetrahydrofluorenyl, or tetrahydroanthracenyl, anthracenyl) ring systemsin which the monocyclic ring system is aromatic or at least one of therings in a bicyclic or tricyclic ring system is aromatic. The bicyclicand tricyclic groups include benzofused 2-3 membered carbocyclic rings.For example, a benzofused group includes phenyl fused with two or moreC₄₋₈ carbocyclic moieties. An aryl is optionally substituted with one ormore substituents including aliphatic [e.g., alkyl, alkenyl, oralkynyl]; cycloaliphatic; (cycloaliphatic)aliphatic;heterocycloaliphatic; (heterocycloaliphatic)aliphatic; aryl; heteroaryl;alkoxy; (cycloaliphatic)oxy; (heterocycloaliphatic)oxy; aryloxy;heteroaryloxy; (araliphatic)oxy; (heteroaraliphatic)oxy; aroyl;heteroaroyl; amino; oxo (on a non-aromatic carbocyclic ring of abenzofused bicyclic or tricyclic aryl); nitro; carboxy; amido; acyl[e.g., (aliphatic)carbonyl; (cycloaliphatic)carbonyl;((cycloaliphatic)aliphatic)carbonyl; (araliphatic)carbonyl;(heterocycloaliphatic)carbonyl;((heterocycloaliphatic)aliphatic)carbonyl; or(heteroaraliphatic)carbonyl]; sulfonyl [e.g., aliphatic-SO₂- oramino-SO₂-]; sulfinyl [e.g., aliphatic-S(O)- or cycloaliphatic-S(O)-];sulfanyl [e.g., aliphatic-S-]; cyano; halo; hydroxy; mercapto; sulfoxy;urea; thiourea; sulfamoyl; sulfamide; or carbamoyl. Alternatively, anaryl can be unsubstituted.

Non-limiting examples of substituted aryls include haloaryl [e.g.,mono-, di (such as

p,m-dihaloaryl), and (trihalo)aryl]; (carboxy)aryl [e.g.,(alkoxycarbonyl)aryl, ((aralkyl)carbonyloxy)aryl, and(alkoxycarbonyl)aryl]; (amido)aryl [e.g., (aminocarbonyl)aryl,(((alkylamino)alkyl)aminocarbonyl)aryl, (alkylcarbonyl)aminoaryl,(arylaminocarbonyl)aryl, and (((heteroaryl)amino)carbonyl)aryl];aminoaryl [e.g., ((alkylsulfonyl)amino)aryl or ((dialkyl)amino)aryl];(cyanoalkyl)aryl; (alkoxy)aryl; (sulfamoyl)aryl [e.g.,(aminosulfonyl)aryl]; (alkylsulfonyl)aryl; (cyano)aryl;(hydroxyalkyl)aryl; ((alkoxy)alkyl)aryl; (hydroxy)aryl,((carboxy)alkyl)aryl; (((dialkyl)amino)alkyl)aryl; (nitroalkyl)aryl;(((alkylsulfonyl)amino)alkyl)aryl; ((heterocycloaliphatic)carbonyl)aryl;((alkylsulfonyl)alkyl)aryl; (cyanoalkyl)aryl; (hydroxyalkyl)aryl;(alkylcarbonyl)aryl; alkylaryl; (trihaloalkyl)aryl;p-amino-m-alkoxycarbonylaryl; p-amino-m-cyanoaryl; p-halo-m-aminoaryl;or (m-(heterocycloaliphatic)-o-(alkyl))aryl.

As used herein, an “araliphatic” such as an “aralkyl” group refers to analiphatic group (e.g., a C₁₋₄ alkyl group) that is substituted with anaryl group. “Aliphatic,” “alkyl,” and “aryl” are defined herein. Anexample of an araliphatic such as an aralkyl group is benzyl.

As used herein, an “aralkyl” group refers to an alkyl group (e.g., aC₁₋₄ alkyl group) that is substituted with an aryl group. Both “alkyl”and “aryl” have been defined above. An example of an aralkyl group isbenzyl. An aralkyl is optionally substituted with one or moresubstituents such as aliphatic [e.g., alkyl, alkenyl, or alkynyl,including carboxyalkyl, hydroxyalkyl, or haloalkyl such astrifluoromethyl], cycloaliphatic [e.g., cycloalkyl or cycloalkenyl],(cycloalkyl)alkyl, heterocycloalkyl, (heterocycloalkyl)alkyl, aryl,heteroaryl, alkoxy, cycloalkyloxy, heterocycloalkyloxy, aryloxy,heteroaryloxy, aralkyloxy, heteroaralkyloxy, aroyl, heteroaroyl, nitro,carboxy, alkoxycarbonyl, alkylcarbonyloxy, amido [e.g., aminocarbonyl,alkylcarbonylamino, cycloalkylcarbonylamino,(cycloalkylalkyl)carbonylamino, arylcarbonylamino, aralkylcarbonylamino,(heterocycloalkyl)carbonylamino, (heterocycloalkylalkyl)carbonylamino,heteroarylcarbonylamino, or heteroaralkylcarbonylamino], cyano, halo,hydroxy, acyl, mercapto, alkylsulfanyl, sulfoxy, urea, thiourea,sulfamoyl, sulfamide, oxo, or carbamoyl.

As used herein, a “bicyclic ring system” includes 8-12 (e.g., 9, 10, or11) membered structures that form two rings, wherein the two rings haveat least one atom in common (e.g., 2 atoms in common). Bicyclic ringsystems include bicycloaliphatics (e.g., bicycloalkyl orbicycloalkenyl), bicycloheteroaliphatics, bicyclic aryls, and bicyclicheteroaryls.

As used herein, a “carbocycle” or “cycloaliphatic” group encompasses a“cycloalkyl” group and a “cycloalkenyl” group, each of which beingoptionally substituted as set forth below.

As used herein, a “cycloalkyl” group refers to a saturated carbocyclicmono- or bicyclic (fused or bridged) ring of 3-10 (e.g., 5-10) carbonatoms. Examples of cycloalkyl groups include cyclopropyl, cyclobutyl,cyclopentyl, cyclohexyl, cycloheptyl, adamantyl, norbornyl, cubyl,octahydro-indenyl, decahydro-naphthyl, bicyclo[3.2.1]octyl,bicyclo[2.2.2]octyl, bicyclo[3.3.1]nonyl, bicyclo[3.3.2.]decyl,bicyclo[2.2.2]octyl, adamantyl, or((aminocarbonyl)cycloalkyl)cycloalkyl.

A “cycloalkenyl” group, as used herein, refers to a non-aromaticcarbocyclic ring of 3-10 (e.g., 4-8) carbon atoms having one or moredouble bonds. Examples of cycloalkenyl groups include cyclopentenyl,1,4-cyclohexa-di-enyl, cycloheptenyl, cyclooctenyl, hexahydro-indenyl,octahydro-naphthyl, cyclohexenyl, cyclopentenyl, bicyclo[2.2.2]octenyl,or bicyclo[3.3.1]nonenyl.

A cycloalkyl or cycloalkenyl group can be optionally substituted withone or more substituents such as phosphor, aliphatic [e.g., alkyl,alkenyl, or alkynyl], cycloaliphatic, (cycloaliphatic) aliphatic,heterocycloaliphatic, (heterocycloaliphatic) aliphatic, aryl,heteroaryl, alkoxy, (cycloaliphatic)oxy, (heterocycloaliphatic)oxy,aryloxy, heteroaryloxy, (araliphatic)oxy, (heteroaraliphatic)oxy, aroyl,heteroaroyl, amino, amido [e.g., (aliphatic)carbonylamino,(cycloaliphatic)carbonylamino, ((cycloaliphatic)aliphatic)carbonylamino,(aryl)carbonylamino, (araliphatic)carbonylamino,(heterocycloaliphatic)carbonylamino,((heterocycloaliphatic)aliphatic)carbonylamino,(heteroaryl)carbonylamino, or (heteroaraliphatic)carbonylamino], nitro,carboxy [e.g., HOOC—, alkoxycarbonyl, or alkylcarbonyloxy], acyl [e.g.,(cycloaliphatic)carbonyl, ((cycloaliphatic) aliphatic)carbonyl,(araliphatic)carbonyl, (heterocycloaliphatic)carbonyl,((heterocycloaliphatic)aliphatic)carbonyl, or(heteroaraliphatic)carbonyl], cyano, halo, hydroxy, mercapto, sulfonyl[e.g., alkyl-SO₂— and aryl-SO₂—], sulfinyl [e.g., alkyl-S(O)—], sulfanyl[e.g., alkyl-S—], sulfoxy, urea, thiourea, sulfamoyl, sulfamide, oxo, orcarbamoyl.

As used herein, the term “heterocycle” or “heterocycloaliphatic”encompasses a heterocycloalkyl group and a heterocycloalkenyl group,each of which being optionally substituted as set forth below.

As used herein, a “heterocycloalkyl” group refers to a 3-10 memberedmono- or bicylic (fused or bridged) (e.g., 5- to 10-membered mono- orbicyclic) saturated ring structure, in which one or more of the ringatoms is a heteroatom (e.g., N, O, S, or combinations thereof). Examplesof a heterocycloalkyl group include piperidyl, piperazyl,tetrahydropyranyl, tetrahydrofuryl, 1,4-dioxolanyl, 1,4-dithianyl,1,3-dioxolanyl, oxazolidyl, isoxazolidyl, morpholinyl, thiomorpholyl,octahydrobenzofuryl, octahydrochromenyl, octahydrothiochromenyl,octahydroindolyl, octahydropyrindinyl, decahydroquinolinyl,octahydrobenzo[b]thiopheneyl, 2-oxa-bicyclo[2.2.2]octyl,1-aza-bicyclo[2.2.2]octyl, 3-aza-bicyclo[3.2.1]octyl, and2,6-dioxa-tricyclo[3.3.1.0^(3,7)]nonyl. A monocyclic heterocycloalkylgroup can be fused with a phenyl moiety to form structures, such astetrahydroisoquinoline, which would be categorized as heteroaryls.

A “heterocycloalkenyl” group, as used herein, refers to a mono- orbicylic (e.g., 5- to 10-membered mono- or bicyclic) non-aromatic ringstructure having one or more double bonds, and wherein one or more ofthe ring atoms is a heteroatom (e.g., N, O, or S). Monocyclic andbicyclic heterocycloaliphatics are numbered according to standardchemical nomenclature.

A heterocycloalkyl or heterocycloalkenyl group can be optionallysubstituted with one or more substituents such as phosphor, aliphatic[e.g., alkyl, alkenyl, or alkynyl], cycloaliphatic,(cycloaliphatic)aliphatic, heterocycloaliphatic,(heterocycloaliphatic)aliphatic, aryl, heteroaryl, alkoxy,(cycloaliphatic)oxy, (heterocycloaliphatic)oxy, aryloxy, heteroaryloxy,(araliphatic)oxy, (heteroaraliphatic)oxy, aroyl, heteroaroyl, amino,amido [e.g., (aliphatic)carbonylamino, (cycloaliphatic)carbonylamino,((cycloaliphatic) aliphatic)carbonylamino, (aryl)carbonylamino,(araliphatic)carbonylamino, (heterocycloaliphatic)carbonylamino,((heterocycloaliphatic) aliphatic)carbonylamino,(heteroaryl)carbonylamino, or (heteroaraliphatic)carbonylamino], nitro,carboxy [e.g., HOOC—, alkoxycarbonyl, or alkylcarbonyloxy], acyl [e.g.,(cycloaliphatic)carbonyl, ((cycloaliphatic) aliphatic)carbonyl,(araliphatic)carbonyl, (heterocycloaliphatic)carbonyl,((heterocycloaliphatic)aliphatic)carbonyl, or(heteroaraliphatic)carbonyl], nitro, cyano, halo, hydroxy, mercapto,sulfonyl [e.g., alkylsulfonyl or arylsulfonyl], sulfinyl [e.g.,alkylsulfinyl], sulfanyl [e.g., alkylsulfanyl], sulfoxy, urea, thiourea,sulfamoyl, sulfamide, oxo, or carbamoyl.

A “heteroaryl” group, as used herein, refers to a monocyclic, bicyclic,or tricyclic ring system having 4 to 15 ring atoms wherein one or moreof the ring atoms is a heteroatom (e.g., N, O, S, or combinationsthereof) and in which the monocyclic ring system is aromatic or at leastone of the rings in the bicyclic or tricyclic ring systems is aromatic.A heteroaryl group includes a benzofused ring system having 2 to 3rings. For example, a benzofused group includes benzo fused with one ortwo 4 to 8 membered heterocycloaliphatic moieties (e.g., indolizyl,indolyl, isoindolyl, 3H-indolyl, indolinyl, benzo[b]furyl,benzo[b]thiophenyl, quinolinyl, or isoquinolinyl). Some examples ofheteroaryl are azetidinyl, pyridyl, 1H-indazolyl, furyl, pyrrolyl,thienyl, thiazolyl, oxazolyl, imidazolyl, tetrazolyl, benzofuryl,isoquinolinyl, benzthiazolyl, xanthene, thioxanthene, phenothiazine,dihydroindole, benzo[1,3]dioxole, benzo[b]furyl, benzo[b]thiophenyl,indazolyl, benzimidazolyl, benzthiazolyl, puryl, cinnolyl, quinolyl,quinazolyl, cinnolyl, phthalazyl, quinazolyl, quinoxalyl, isoquinolyl,4H-quinolizyl, benzo-1,2,5-thiadiazolyl, or 1,8-naphthyridyl.

Without limitation, monocyclic heteroaryls include furyl, thiophenyl,2H-pyrrolyl, pyrrolyl, oxazolyl, thazolyl, imidazolyl, pyrazolyl,isoxazolyl, isothiazolyl, 1,3,4-thiadiazolyl, 2H-pyranyl, 4-H-pranyl,pyridyl, pyridazyl, pyrimidyl, pyrazolyl, pyrazyl, or 1,3,5-triazyl.Monocyclic heteroaryls are numbered according to standard chemicalnomenclature.

Without limitation, bicyclic heteroaryls include indolizyl, indolyl,isoindolyl, 3H-indolyl, indolinyl, benzo[b]furyl, benzo[b]thiophenyl,quinolinyl, isoquinolinyl, indolizinyl, isoindolyl, indolyl,benzo[b]furyl, bexo[b]thiophenyl, indazolyl, benzimidazyl,benzthiazolyl, purinyl, 4H-quinolizyl, quinolyl, isoquinolyl, cinnolyl,phthalazyl, quinazolyl, quinoxalyl, 1,8-naphthyridyl, or pteridyl.Bicyclic heteroaryls are numbered according to standard chemicalnomenclature.

A heteroaryl is optionally substituted with one or more substituentssuch as aliphatic [e.g., alkyl, alkenyl, or alkynyl]; cycloaliphatic;(cycloaliphatic)aliphatic; heterocycloaliphatic;(heterocycloaliphatic)aliphatic; aryl; heteroaryl; alkoxy;(cycloaliphatic)oxy; (heterocycloaliphatic)oxy; aryloxy; heteroaryloxy;(araliphatic)oxy; (heteroaraliphatic)oxy; aroyl; heteroaroyl; amino; oxo(on a non-aromatic carbocyclic or heterocyclic ring of a bicyclic ortricyclic heteroaryl); carboxy; amido; acyl [e.g., aliphaticcarbonyl;(cycloaliphatic)carbonyl; ((cycloaliphatic)aliphatic)carbonyl;(araliphatic)carbonyl; (heterocycloaliphatic)carbonyl;((heterocycloaliphatic)aliphatic)carbonyl; or(heteroaraliphatic)carbonyl]; sulfonyl [e.g., aliphaticsulfonyl oraminosulfonyl]; sulfinyl [e.g., aliphaticsulfinyl]; sulfanyl [e.g.,aliphaticsulfanyl]; nitro; cyano; halo; hydroxy; mercapto; sulfoxy;urea; thiourea; sulfamoyl; sulfamide; or carbamoyl. Alternatively, aheteroaryl can be unsubstituted.

Non-limiting examples of substituted heteroaryls include(halo)heteroaryl [e.g., mono- and di-(halo)heteroaryl];(carboxy)heteroaryl [e.g., (alkoxycarbonyl)heteroaryl]; cyanoheteroaryl;aminoheteroaryl [e.g., ((alkylsulfonyl)amino)heteroaryl and((dialkyl)amino)heteroaryl]; (amido)heteroaryl [e.g.,aminocarbonylheteroaryl, ((alkylcarbonyl)amino)heteroaryl,((((alkyl)amino)alkyl)aminocarbonyl)heteroaryl,(((heteroaryl)amino)carbonyl)heteroaryl,((heterocycloaliphatic)carbonyl)heteroaryl, and((alkylcarbonyl)amino)heteroaryl]; (cyanoalkyl)heteroaryl;(alkoxy)heteroaryl; (sulfamoyl)heteroaryl [e.g.,(aminosulfonyl)heteroaryl]; (sulfonyl)heteroaryl [e.g.,(alkylsulfonyl)heteroaryl]; (hydroxyalkyl)heteroaryl;(alkoxyalkyl)heteroaryl; (hydroxy)heteroaryl;((carboxy)alkyl)heteroaryl; (((dialkyl)amino)alkyl]heteroaryl;(heterocycloaliphatic)heteroaryl; (cycloaliphatic)heteroaryl;(nitroalkyl)heteroaryl; (((alkylsulfonyl)amino)alkyl)heteroaryl;((alkylsulfonyl)alkyl)heteroaryl; (cyanoalkyl)heteroaryl;(acyl)heteroaryl [e.g., (alkylcarbonyl)heteroaryl]; (alkyl)heteroaryl,and (haloalkyl)heteroaryl [e.g., trihaloalkylheteroaryl].

A “heteroaraliphatic” (such as a heteroaralkyl group) as used herein,refers to an aliphatic group (e.g., a C₁₋₄ alkyl group) that issubstituted with a heteroaryl group. “Aliphatic,” “alkyl,” and“heteroaryl” have been defined above.

A “heteroaralkyl” group, as used herein, refers to an alkyl group (e.g.,a C₁₋₄ alkyl group) that is substituted with a heteroaryl group. Both“alkyl” and “heteroaryl” have been defined above. A heteroaralkyl isoptionally substituted with one or more substituents such as alkyl(including carboxyalkyl, hydroxyalkyl, and haloalkyl such astrifluoromethyl), alkenyl, alkynyl, cycloalkyl, (cycloalkyl)alkyl,heterocycloalkyl, (heterocycloalkyl)alkyl, aryl, heteroaryl, alkoxy,cycloalkyloxy, heterocycloalkyloxy, aryloxy, heteroaryloxy, aralkyloxy,heteroaralkyloxy, aroyl, heteroaroyl, nitro, carboxy, alkoxycarbonyl,alkylcarbonyloxy, aminocarbonyl, alkylcarbonylamino,cycloalkylcarbonylamino, (cycloalkylalkyl)carbonylamino,arylcarbonylamino, aralkylcarbonylamino,(heterocycloalkyl)carbonylamino, (heterocycloalkylalkyl)carbonylamino,heteroarylcarbonylamino, heteroaralkylcarbonylamino, cyano, halo,hydroxy, acyl, mercapto, alkylsulfanyl, sulfoxy, urea, thiourea,sulfamoyl, sulfamide, oxo, or carbamoyl.

As used herein, “cyclic moiety” and “cyclic group” refer to mono-, bi-,and tri-cyclic ring systems including cycloaliphatic,heterocycloaliphatic, aryl, or heteroaryl, each of which has beenpreviously defined.

As used herein, a “bridged bicyclic ring system” refers to a bicyclicheterocyclicaliphatic ring system or bicyclic cycloaliphatic ring systemin which the rings are bridged. Examples of bridged bicyclic ringsystems include, but are not limited to, adamantanyl, norbomanyl,bicyclo[3.2.1]octyl, bicyclo[2.2.2]octyl, bicyclo[3.3.1]nonyl,bicyclo[3.2.3]nonyl, 2-oxabicyclo[2.2.2]octyl, 1-azabicyclo[2.2.2]octyl,3-azabicyclo[3.2.1]octyl, and 2,6-dioxa-tricyclo[3.3.1.0^(3,7)]nonyl. Abridged bicyclic ring system can be optionally substituted with one ormore substituents such as alkyl (including carboxyalkyl, hydroxyalkyl,and haloalkyl such as trifluoromethyl), alkenyl, alkynyl, cycloalkyl,(cycloalkyl)alkyl, heterocycloalkyl, (heterocycloalkyl)alkyl, aryl,heteroaryl, alkoxy, cycloalkyloxy, heterocycloalkyloxy, aryloxy,heteroaryloxy, aralkyloxy, heteroaralkyloxy, aroyl, heteroaroyl, nitro,carboxy, alkoxycarbonyl, alkylcarbonyloxy, aminocarbonyl,alkylcarbonylamino, cycloalkylcarbonylamino,(cycloalkylalkyl)carbonylamino, arylcarbonylamino, aralkylcarbonylamino,(heterocycloalkyl)carbonylamino, (heterocycloalkylalkyl)carbonylamino,heteroarylcarbonylamino, heteroaralkylcarbonylamino, cyano, halo,hydroxy, acyl, mercapto, alkylsulfanyl, sulfoxy, urea, thiourea,sulfamoyl, sulfamide, oxo, or carbamoyl.

As used herein, an “acyl” group refers to a formyl group or R^(X)—C(O)—(such as alkyl-C(O)—, also referred to as “alkylcarbonyl”) where R^(X)and “alkyl” have been defined previously. Acetyl and pivaloyl areexamples of acyl groups.

As used herein, an “aroyl” or “heteroaroyl” refers to an aryl-C(O)- or aheteroaryl-C(O)—. The aryl and heteroaryl portion of the aroyl orheteroaroyl is optionally substituted as previously defined.

As used herein, an “alkoxy” group refers to an alkyl-O— group where“alkyl” has been defined previously.

As used herein, a “carbamoyl” group refers to a group having thestructure —O—CO—NR^(X)R^(Y) or —NR^(X)—CO—O—R^(Z), wherein R^(X) andR^(Y) have been defined above and R^(Z) can be aliphatic, aryl,araliphatic, heterocycloaliphatic, heteroaryl, or heteroaraliphatic.

As used herein, a “carboxy” group refers to —COOH, —COOR^(X), —OC(O)H,—OC(O)R^(X), when used as a terminal group; or —OC(O)— or —C(O)O— whenused as an internal group.

As used herein, a “haloaliphatic” group refers to an aliphatic groupsubstituted with 1-3 halogen. For instance, the term haloalkyl includesthe group —CF₃.

As used herein, a “mercapto” group refers to —SH.

As used herein, a “sulfo” group refers to —SO₃H or —SO₃R^(X) when usedterminally or —S(O)₃— when used internally.

As used herein, a “sulfamide” group refers to the structure—NR^(X)—S(O)₂—NR^(Y)R^(Z) when used terminally and —NR^(X)—S(O)₂—NR^(Y)—when used internally, wherein R^(X), R^(Y), and R^(Z) have been definedabove.

As used herein, a “sulfonamide” group refers to the structure—S(O)₂—NR^(X)R^(Y) or —NR^(X)—S(O)₂—R^(Z) when used terminally; or—S(O)₂—NR^(X)— or —NR^(X)—S(O)₂— when used internally, wherein R^(X),R^(Y), and R^(Z) are defined above.

As used herein a “sulfanyl” group refers to —S—R^(X) when usedterminally and —S— when used internally, wherein R^(X) has been definedabove. Examples of sulfanyls include aliphatic-S—, cycloaliphatic-S—,aryl-S—, or the like.

As used herein a “sulfinyl” group refers to —S(O)—R^(X) when usedterminally and —S(O)— when used internally, wherein R^(X) has beendefined above. Exemplary sulfinyl groups include aliphatic-S(O)—,aryl-S(O)—, (cycloaliphatic(aliphatic))—S(O)—, cycloalkyl-S(O)—,heterocycloaliphatic-S(O)—, heteroaryl-S(O)—, or the like.

As used herein, a “sulfonyl” group refers to —S(O)₂—R^(X) when usedterminally and —S(O)₂— when used internally, wherein R^(X) has beendefined above. Exemplary sulfonyl groups include aliphatic-S(O)₂—,aryl-S(O)₂—, (cycloaliphatic(aliphatic))—S(O)₂—, cycloaliphatic-S(O)₂—,heterocycloaliphatic-S(O)₂—, heteroaryl-S(O)₂—,(cycloaliphatic(amido(aliphatic)))—S(O)₂— or the like.

As used herein, a “sulfoxy” group refers to —O—SO—R^(X) or —SO—O—R^(X),when used terminally and —O—S(O)— or —S(O)—O— when used internally,where R^(X) has been defined above.

As used herein, a “halogen” or “halo” group refers to fluorine,chlorine, bromine or iodine.

As used herein, an “alkoxycarbonyl,” which is encompassed by the termcarboxy, used alone or in connection with another group refers to agroup such as alkyl-O—C(O)—.

As used herein, an “alkoxyalkyl” refers to an alkyl group such asalkyl-O-alkyl-, wherein alkyl has been defined above.

As used herein, a “carbonyl” refer to —C(O)—.

As used herein, an “oxo” refers to ═O.

As used herein, the term “phospho” refers to phosphinates andphosphonates. Examples of phosphinates and phosphonates include—P(O)(R^(P))₂, wherein R^(P) is aliphatic, alkoxy, aryloxy,heteroaryloxy, (cycloaliphatic)oxy, (heterocycloaliphatic)oxy aryl,heteroaryl, cycloaliphatic or amino.

As used herein, an “aminoalkyl” refers to the structure(R^(X))₂N-alkyl-.

As used herein, a “cyanoalkyl” refers to the structure (NC)-alkyl-.

As used herein, a “urea” group refers to the structure—NR^(X)—CO—NR^(Y)R^(Z) and a “thiourea” group refers to the structure—NR^(X)—CS—NR^(Y)R^(Z) when used terminally and —NR^(X)—CO—NR^(Y)— or—NR^(X)—CS—NR^(Y)— when used internally, wherein R^(X), R^(Y), and R^(Z)have been defined above.

As used herein, a “guanidine” group refers to the structure—N═C(N(R^(X)R^(Y)))N(R^(X)R^(Y)) or —NR^(X)—C(═NR^(X))NR^(X)R^(Y)wherein R^(X) and R^(Y) have been defined above.

As used herein, the term “amidino” group refers to the structure—C═(NR^(X))N(R^(X)R^(Y)) wherein R^(X) and R^(Y) have been definedabove.

In general, the term “vicinal” refers to the placement of substituentson a group that includes two or more carbon atoms, wherein thesubstituents are attached to adjacent carbon atoms.

In general, the term “geminal” refers to the placement of substituentson a group that includes two or more carbon atoms, wherein thesubstituents are attached to the same carbon atom.

The terms “terminally” and “internally” refer to the location of a groupwithin a substituent. A group is terminal when the group is present atthe end of the substituent not further bonded to the rest of thechemical structure. Carboxyalkyl, i.e., R^(X)O(O)C-alkyl is an exampleof a carboxy group used terminally. A group is internal when the groupis present in the middle of a substituent of the chemical structure.Alkylcarboxy (e.g., alkyl-C(O)O— or alkyl-OC(O)—) and alkylcarboxyaryl(e.g., alkyl-C(O)O-aryl- or alkyl-O(CO)-aryl-) are examples of carboxygroups used internally.

As used herein, an “aliphatic chain” refers to a branched or straightaliphatic group (e.g., alkyl groups, alkenyl groups, or alkynyl groups).A straight aliphatic chain has the structure —[CH₂]_(v)—, where v is1-12. A branched aliphatic chain is a straight aliphatic chain that issubstituted with one or more aliphatic groups. A branched aliphaticchain has the structure —[CQQ]_(v)-where each Q is independently ahydrogen or an aliphatic group; however, Q shall be an aliphatic groupin at least one instance. The term aliphatic chain includes alkylchains, alkenyl chains, and alkynyl chains, where alkyl, alkenyl, andalkynyl are defined above.

The phrase “optionally substituted” is used interchangeably with thephrase “substituted or unsubstituted.” As described herein, compounds ofthe invention can optionally be substituted with one or moresubstituents, such as are illustrated generally above, or as exemplifiedby particular classes, subclasses, and species of the invention. Asdescribed herein, the variables R₁, R₂, and R₃, and other variablescontained in formulae described herein encompass specific groups, suchas alkyl and aryl. Unless otherwise noted, each of the specific groupsfor the variables R₁, R₂, and R₃, and other variables contained thereincan be optionally substituted with one or more substituents describedherein. Each substituent of a specific group is further optionallysubstituted with one to three of halo, cyano, oxo, alkoxy, hydroxy,amino, nitro, aryl, cycloaliphatic, heterocycloaliphatic, heteroaryl,haloalkyl, and alkyl. For instance, an alkyl group can be substitutedwith alkylsulfanyl and the alkylsulfanyl can be optionally substitutedwith one to three of halo, cyano, oxo, alkoxy, hydroxy, amino, nitro,aryl, haloalkyl, and alkyl. As an additional example, the cycloalkylportion of a (cycloalkyl)carbonylamino can be optionally substitutedwith one to three of halo, cyano, alkoxy, hydroxy, nitro, haloalkyl, andalkyl. When two alkoxy groups are bound to the same atom or adjacentatoms, the two alkxoy groups can form a ring together with the atom(s)to which they are bound.

In general, the term “substituted,” whether preceded by the term“optionally” or not, refers to the replacement of hydrogen radicals in agiven structure with the radical of a specified substituent. Specificsubstituents are described above in the definitions and below in thedescription of compounds and examples thereof. Unless otherwiseindicated, an optionally substituted group can have a substituent ateach substitutable position of the group, and when more than oneposition in any given structure can be substituted with more than onesubstituent selected from a specified group, the substituent can beeither the same or different at every position. A ring substituent, suchas a heterocycloalkyl, can be bound to another ring, such as acycloalkyl, to form a spiro-bicyclic ring system, e.g., both rings shareone common atom. As one of ordinary skill in the art will recognize,combinations of substituents envisioned by this invention are thosecombinations that result in the formation of stable or chemicallyfeasible compounds.

The phrase “stable or chemically feasible,” as used herein, refers tocompounds that are not substantially altered when subjected toconditions to allow for their production, detection, and preferablytheir recovery, purification, and use for one or more of the purposesdisclosed herein. In some embodiments, a stable compound or chemicallyfeasible compound is one that is not substantially altered when kept ata temperature of 40° C. or less, in the absence of moisture or otherchemically reactive conditions, for at least a week.

As used herein, an “effective amount” is defined as the amount requiredto confer a therapeutic effect on the treated patient, and is typicallydetermined based on age, surface area, weight, and condition of thepatient. The interrelationship of dosages for animals and humans (basedon milligrams per meter squared of body surface) is described byFreireich et al., Cancer Chemother. Rep., 50: 219 (1966). Body surfacearea may be approximately determined from height and weight of thepatient. See, e.g., Scientific Tables, Geigy Pharmaceuticals, Ardsley,N.Y., 537 (1970). As used herein, “patient” refers to a mammal,including a human.

Unless otherwise stated, structures depicted herein are also meant toinclude all isomeric (e.g., enantiomeric, diastereomeric, and geometric(or conformational)) forms of the structure; for example, the R and Sconfigurations for each asymmetric center, (Z) and (E) double bondisomers, and (Z) and (E) conformational isomers. Therefore, singlestereochemical isomers as well as enantiomeric, diastereomeric, andgeometric (or conformational) mixtures of the present compounds arewithin the scope of the invention. Unless otherwise stated, alltautomeric forms of the compounds of the invention are within the scopeof the invention. Additionally, unless otherwise stated, structuresdepicted herein are also meant to include compounds that differ only inthe presence of one or more isotopically enriched atoms. For example,compounds having the present structures except for the replacement ofhydrogen by deuterium or tritium, or the replacement of a carbon by a¹³C- or ¹⁴C-enriched carbon are within the scope of this invention. Suchcompounds are useful, for example, as analytical tools or probes inbiological assays, or as therapeutic agents.

Compounds of the present invention are useful modulators of ABCtransporters and are useful in the treatment of ABC transporter mediateddiseases.

II. Compounds A. Generic Compounds

The present invention relates to compounds of formula I useful asmodulators of ABC transporter activity:

or a pharmaceutically acceptable salt thereof.

R₁ is —Z^(A)R₄, wherein each Z^(A) is independently a bond or anoptionally substituted branched or straight C₁₋₆ aliphatic chain whereinup to two carbon units of Z^(A) are optionally and independentlyreplaced by —CO—, —CS—, —CONR^(A)—, —CONR^(A)NR^(A)—, —CO₂—, —OCO—,—NR^(A)CO₂-, —O—, —NR^(A)CONR^(A)—, —OCONR^(A)—, —NR^(A)NR^(A)—,—NR^(A)CO—, —S—, —SO—, —SO₂—, —NR^(A)—, —SO₂NR^(A)—; NR^(A)SO₂—, or—NR^(A)SO₂NR^(A)—. Each R₄ is independently R^(A), halo, —OH, —NH₂,—NO₂, —CN, or —OCF₃. Each R^(A) is independently hydrogen, an optionallysubstituted aliphatic, an optionally substituted cycloaliphatic, anoptionally substituted heterocycloaliphatic, an optionally substitutedaryl, or an optionally substituted heteroaryl.

R₂ is —Z^(B)R₅, wherein each Z^(B) is independently a bond or anoptionally substituted branched or straight C₁₋₆ aliphatic chain whereinup to two carbon units of Z^(B) are optionally and independentlyreplaced by —CO—, —CS—, —CONR^(B)—, —CONR^(B)NR^(B)—, —CO₂—, —OCO—,—NR^(B)CO₂—, —O—, —NR^(B)CONR^(B)—, —OCONR^(B)—, —NR^(B)NR^(B)—,—NR^(B)CO—, —S—, —SO—, —SO₂-, —NR^(B)—, —SO₂NR^(B)—, —NR^(B)SO₂—, or—NR^(B)SO₂NR^(B)—. Each R₅ is independently R^(B), halo, —OH, —NH₂,—NO₂, —CN, —CF₃, or —OCF₃. Each R^(B) is independently hydrogen, anoptionally substituted aliphatic, an optionally substitutedcycloaliphatic, an optionally substituted heterocycloaliphatic, anoptionally substituted aryl, or an optionally substituted heteroaryl.Alternatively, any two adjacent R₂ groups together with the atoms towhich they are attached form an optionally substituted carbocycle or anoptionally substituted heterocycle.

Ring A is an optionally substituted 3-7 membered monocyclic ring having0-3 heteroatoms selected from N, O, and S.

Ring B is a group having formula Ia:

or a pharmaceutically acceptable salt thereof, wherein p is 0-3 and eachR₃ and R′₃ is independently —Z^(C)R₆, where each Z^(C) is independentlya bond or an optionally substituted branched or straight C₁₋₆ aliphaticchain wherein up to two carbon units of Z^(C) are optionally andindependently replaced by —CO—, —CS—, —CONR^(C)—, —CONR^(C)NR^(C)—,—CO₂—, —OCO—, —NR^(C)CO₂-, —O—, —NR^(C)CONR^(C)—, —OCONR^(C)—,—NR^(C)NR^(C)—, —NR^(C)CO—, —S—, —SO—, —SO₂—, —NR^(C)—, —SO₂NR^(C)—,—NR^(C)SO₂—, or —NR^(C)SO₂NR^(C)—. Each R₆ is independently R^(C), halo,—OH, —NH₂, —NO₂, —CN, or —OCF₃. Each R^(C) is independently hydrogen, anoptionally substituted aliphatic, an optionally substitutedcycloaliphatic, an optionally substituted heterocycloaliphatic, anoptionally substituted aryl, or an optionally substituted heteroaryl.Alternatively, any two adjacent R₃ groups together with the atoms towhich they are attached form an optionally substituted carbocycle or anoptionally substituted heterocycle. Furthermore, R′₃ and an adjacent R₃group, together with the atoms to which they are attached, form anoptionally substituted heterocycle.

n is 1-3.

However, in several embodiments, when ring A is unsubstitutedcyclopentyl, n is 1, R₂ is 4-chloro, and R₁ is hydrogen, then ring B isnot 2-(tertbutyl)indol-5-yl, or(2,6-dichlorophenyl(carbonyl))-3-methyl-1H-indol-5-yl; and when ring Ais unsubstituted cyclopentyl, n is 0, and R₁ is hydrogen, then ring B isnot

B. Specific Compounds

1. R₁ Group

R₁ is —Z^(A)R₄, wherein each Z^(A) is independently a bond or anoptionally substituted branched or straight C₁₋₆ aliphatic chain whereinup to two carbon units of Z^(A) are optionally and independentlyreplaced by —CO—, —CS—, —CONR^(A)—, —CONR^(A)NR^(A)—, —CO₂—, —OCO—,—NR^(A)CO₂—,

—O—, —NR^(A)CONR^(A)—, —OCONR^(A)—, —NR^(A)NR^(A)—, —NR^(A)CO—, —S—,—SO—, —SO₂—, —NR^(A)—, —SO₂NR^(A)—, —NR^(A)SO₂—, or —NR^(A)SO₂NR^(A)—.Each R₄ is independently R^(A), halo, —OH, —NH₂, —NO₂, —CN, or —OCF₃.Each R^(A) is independently hydrogen, an optionally substitutedaliphatic, an optionally substituted cycloaliphatic, an optionallysubstituted heterocycloaliphatic, an optionally substituted aryl, or anoptionally substituted heteroaryl.

In several embodiments, R₁ is —Z^(A)R₄, wherein each Z^(A) isindependently a bond or an optionally substituted branched or straightC₁₋₆ aliphatic chain and each R₄ is hydrogen.

In other embodiments, R₁ is —Z^(A)R₄, wherein each Z^(A) is a bond andeach R₄ is hydrogen.

2. R₂ Group

Each R₂ is independently —Z^(B)R₅, wherein each Z^(B) is independently abond or an optionally substituted branched or straight C₁₋₆ aliphaticchain wherein up to two carbon units of Z^(B) are optionally andindependently replaced by —CO—, —CS—, —CONR^(B)—, —CONR^(B)NR^(B)——CO₂—,—OCO—, —NR^(B)CO₂—, —O—, —NR^(B)CONR^(B)—, —OCONR^(B)—, —NR^(B)NR^(B)—,—NR^(B)CO—, —S—, —SO—, —SO₂—, —NR^(B)—, —SO₂NR^(B)—, —NR^(B)SO₂—, or—NR^(B)SO₂NR^(B)—. Each R₅ is independently R^(B), halo, —OH, —NH₂,—NO₂, —CN, —CF₃, or —OCF₃. Each R^(B) is independently hydrogen, anoptionally substituted aliphatic, an optionally substitutedcycloaliphatic, an optionally substituted heterocycloaliphatic, anoptionally substituted aryl, or an optionally substituted heteroaryl.Alternatively, any two adjacent R₂ groups together with the atoms towhich they are attached form an optionally substituted carbocycle or anoptionally substituted heterocycle, or an optionally substitutedheteroaryl.

In several embodiments, R₂ is an optionally substituted aliphatic. Forexample, R₂ is an optionally substituted branched or straight C₁₋₆aliphatic chain. In other examples, R₂ is an optionally substitutedbranched or straight C₁₋₆ alkyl chain, an optionally substitutedbranched or straight C₂₋₆ alkenyl chain, or an optionally substitutedbranched or straight C₂₋₆ alkynyl chain. In alternative embodiments, R₂is a branched or straight C₁₋₆ aliphatic chain that is optionallysubstituted with 1-3 of halo, hydroxy, cyano, cycloaliphatic,heterocycloaliphatic, aryl, heteroaryl, or combinations thereof. Forexample, R₂ is a branched or straight C₁₋₆ alkyl that is optionallysubstituted with 1-3 of halo, hydroxy, cyano, cycloaliphatic,heterocycloaliphatic, aryl, heteroaryl, or combinations thereof. Instill other examples, R₂ is a methyl, ethyl, propyl, butyl, isopropyl,or tert-butyl, each of which is optionally substituted with 1-3 of halo,hydroxy, cyano, aryl, heteroaryl, cycloaliphatic, orheterocycloaliphatic. In still other examples, R₂ is a methyl, ethyl,propyl, butyl, isopropyl, or tert-butyl, each of which is unsubstituted.

In several other embodiments, R₂ is an optionally substituted branchedor straight C₁₋₅ alkoxy. For example, R₂ is a C1.5 alkoxy that isoptionally substituted with 1-3 of hydroxy, aryl, heteroaryl,cycloaliphatic, heterocycloaliphatic, or combinations thereof. In otherexamples, R₂ is a methoxy, ethoxy, propoxy, butoxy, or pentoxy, each ofwhich is optionally substituted with 1-3 of hydroxy, aryl, heteroaryl,cycloaliphatic, heterocycloaliphatic, or combinations thereof.

In other embodiments, R₂ is hydroxy, halo, or cyano.

In several embodiments, R₂ is —Z^(B)R₅, and Z^(B) is independently abond or an optionally substituted branched or straight C₁₋₄ aliphaticchain wherein up to two carbon units of Z^(B) are optionally andindependently replaced by —C(O)—, —O—, —S—, —S(O)₂—, or —NH—, and R₅ isR^(B), halo, —OH, —NH₂, —NO₂, —CN, —CF₃, or —OCF₃, and R^(B) is hydrogenor aryl.

In several embodiments, two adjacent R₂ groups form an optionallysubstituted carbocycle or an optionally substituted heterocycle. Forexample, two adjacent R₂ groups form an optionally substitutedcarbocycle or an optionally substituted heterocycle, either of which isfused to the phenyl of formula I, wherein the carbocycle or heterocyclehas formula Ib:

Each of Z₁, Z₂, Z₃, Z₄, and Z₅ is independently a bond, —CR₇R′₇—,—C(O)—, —NR₇—, or —O—; each R₇ is independently —Z^(D)R₈, wherein eachZ^(D) is independently a bind or an optionally substituted branched orstraight C₁₋₆ aliphatic chain wherein up to two carbon units of Z^(D)are optionally and independently replaced by —CO—, —CS—, —CONR^(D)—,—CO₂—, —OCO—, —NR^(D)CO₂—, —O—,

—NR^(D)CONR^(D)—, —OCONR^(D)—, —NR^(D)NR^(D)—, —NR^(D)CO—, —S—, —SO—,—SO₂—, —NR^(D)—, —SO₂NR^(D) NR^(D)SO₂—, or —NR^(D)SO₂NR^(D)—. Each R₈ isindependently RD, halo, —OH, —NH₂, —NO₂, —CN, —CF₃, or —OCF₃. Each R^(D)is independently hydrogen, an optionally substituted cycloaliphatic, anoptionally substituted heterocycloaliphatic, an optionally substitutedaryl, or an optionally substituted heteroaryl. Each R′₇ is independentlyhydrogen, optionally substituted C₁₋₆ aliphatic, hydroxy, halo, cyano,nitro, or combinations thereof. Alternatively, any two adjacent R₇groups together with the atoms to which they are attached form anoptionally substituted 3-7 membered carbocyclic ring, such as anoptionally substituted cyclobutyl ring, or any two R₇ and R′₇ groupstogether with the atom or atoms to which they are attached form anoptionally substituted 3-7 membered carbocyclic ring or aheterocarbocyclic ring.

In several other examples, two adjacent R₂ groups form an optionallysubstituted carbocycle. For example, two adjacent R₂ groups form anoptionally substituted 5-7 membered carbocycle that is optionallysubstituted with 1-3 of halo, hydroxy, cyano, oxo, cyano, alkoxy, alkyl,or combinations thereof. In another example, two adjacent R₂ groups forma 5-6 membered carbocycle that is optionally substituted with 1-3 ofhalo, hydroxy, cyano, oxo, cyano, alkoxy, alkyl, or combinationsthereof. In still another example, two adjacent R₂ groups form anunsubstituted 5-7 membered carbocycle.

In alternative examples, two adjacent R₂ groups form an optionallysubstituted heterocycle. For instance, two adjacent R₂ groups form anoptionally substituted 5-7 membered heterocycle having 1-3 heteroatomsindependently selected from N, O, and S. In several examples, twoadjacent R₂ groups form an optionally substituted 5-6 memberedheterocycle having 1-2 oxygen atoms. In other examples, two adjacent R₂groups form an unsubstituted 5-7 membered heterocycle having 1-2 oxygenatoms. In other embodiments, two adjacent R₂ groups form a ring selectedfrom:

In alternative examples, two adjacent R₂ groups form an optionallysubstituted carbocycle or an optionally substituted heterocycle, and athird R₂ group is attached to any chemically feasible position on thephenyl of formula I. For instance, an optionally substituted carbocycleor an optionally substituted heterocycle, both of which is formed by twoadjacent R₂ groups; a third R₂ group; and the phenyl of formula I form agroup having formula Ic:

Z₁, Z₂, Z₃, Z₄, and Z₅ has been defined above in formula Ib, and R₂ hasbeen defined above in formula I.

In several embodiments, each R₂ group is independently selected fromhydrogen, halo,

—OCH₃, —OH, —CH₂OH, —CH₃, and —OCF₃, and/or two adjacent R₂ groupstogether with the atoms to which they are attached form

In other embodiments, R₂ is at least one selected from hydrogen, halo,methoxy, phenylmethoxy, hydroxy, hydroxymethyl, trifluoromethoxy, andmethyl.

In some embodiments, two adjacent R₂ groups, together with the atoms towhich they are attached, form

3. Ring A

Ring A is an optionally substituted 3-7 membered monocyclic ring having0-3 heteroatoms selected from N, O, and S.

In several embodiments, ring A is an optionally substituted 3-7 memberedmonocyclic cycloaliphatic. For example, ring A is a cyclopropyl,cyclobutyl, cyclopentyl, cyclohexyl, or cycloheptyl, each of which isoptionally substituted with 1-3 of halo, hydroxy, C₁₋₅ aliphatic, orcombinations thereof.

In other embodiments, ring A is an optionally substituted 3-7 memberedmonocyclic heterocycloaliphatic. For example, ring A is an optionallysubstituted 3-7 membered monocyclic heterocycloaliphatic having 1-2heteroatoms independently selected from N, O, and S. In other examples,ring A is tetrahydrofuran-yl, tetrahydro-2H-pyran-yl, pyrrolidone-yl, orpiperidine-yl, each of which is optionally substituted.

In still other examples, ring A is selected from

Each R₈ is independently —Z^(E)R₉, wherein each Z^(E) is independently abond or an optionally substituted branched or straight C₁₋₅ aliphaticchain wherein up to two carbon units of Z^(E) are optionally andindependently replaced by —CO—, —CS—, —CONR^(E)—, —C₂—, —OCO—,—NR^(E)CO₂—, —O—, —NR^(E)CONR^(E)—, —OCONR^(E)—, —NR^(E)NR^(E)—,—NR^(E)CO—, —S—, —SO—, —SO₂—, —NR^(E)—, —SO₂NR^(E)—, —NR^(E)SO₂—, or—NR^(E)SO₂NR^(E)—, each R₉ is independently R^(E), —OH, —NH₂, —NO₂, —CN,—CF₃, oxo, or —OCF₃. Each R^(E) is independently hydrogen, an optionallysubstituted cycloaliphatic, an optionally substitutedheterocycloaliphatic, an optionally substituted aryl, or an optionallysubstituted heteroaryl.

q is 0-5.

In other embodiments, ring A is one selected from

In several embodiments, ring A is

4. Ring B

Ring B is a group having formula Ia:

or a pharmaceutically acceptable salt thereof, wherein p is 0-3.

Each R₃ and R′₃ is independently —Z^(C)R₆, where each Z^(C) isindependently a bond or an optionally substituted branched or straightC₁₋₆ aliphatic chain wherein up to two carbon units of Z^(C) areoptionally and independently replaced by —CO—, —CS—, —CONR^(C)—,—CONR^(C)NR^(C)—, —CO₂—, —OCO—, —NR^(C)CO₂—, —O—, —NR^(C)CONR^(C)—,—OCONR^(C)—, —NR^(C)NR^(C)—, —NR^(C)CO—, —S—, —SO—, —SO₂—, —NR^(C)—,—SO₂NR^(C)—, —NR^(C)SO₂—, or —NR^(C)SO₂NR^(C)—. Each R₆ is independentlyR^(C), halo, —OH, —NH₂, —NO₂, —CN, or —OCF₃. Each R^(C) is independentlyhydrogen, an optionally substituted aliphatic, an optionally substitutedcycloaliphatic, an optionally substituted heterocycloaliphatic, anoptionally substituted aryl, or an optionally substituted heteroaryl.Alternatively, any two adjacent R₃ groups together with the atoms towhich they are attached form an optionally substituted carbocycle or anoptionally substituted heterocycle, or R′₃ and an adjacent R₃, i.e.,attached to the 2 position of the indole of formula Ia, together withthe atoms to which they are attached form an optionally substitutedheterocycle.

In several embodiments, ring B is

wherein q is 0-3 and each R₂₀ is —Z^(G)R₂₁, where each Z^(G) isindependently a bond or an optionally substituted branched or straightC₁₋₅ aliphatic chain wherein up to two carbon units of Z^(G) areoptionally and independently replaced by —CO—, —CS—, —CONR^(G), —CO2—,—OCO—, —NR^(G)CO₂—, —O—, —OCONR^(G)—, —NR^(G)NR^(G)—, —NR^(G)CO—, —S—,—SO—, —SO₂—, —NR^(G)—, —SO₂NR^(G)—, —NR^(G)SO₂—, or —NR^(G)SO₂NR^(G)—.Each R₂₁ is independently R^(G), halo, —OH, —NH₂, —NO₂, —CN, or —OCF₃.Each R^(G) is independently hydrogen, an optionally substitutedaliphatic, an optionally substituted cycloaliphatic, an optionallysubstituted heterocycloaliphatic, an optionally substituted aryl, or anoptionally substituted heteroaryl.

For example, ring B is

In several embodiments, R′₃ is hydrogen and R₃ is attached to the 2, 3,4, 5, 6, or 7 position of the indole of formula Ia. In several otherexamples, R₃ is attached to the 2 or 3 position of the indole of formulaIa, and R₃ is independently an optionally substituted aliphatic. Forinstance, R₃ is an optionally substituted acyl group. In severalinstances, R₃ is an optionally substituted (alkoxy)carbonyl. In otherinstances, R₃ is (methoxy)carbonyl, (ethoxy)carbonyl, (propoxy)carbonyl,or (butoxy)carbonyl, each of which is optionally substituted with 1-3 ofhalo, hydroxy, or combinations thereof. In other instances, R₃ is anoptionally substituted (aliphatic)carbonyl. For example, R₃ is anoptionally substituted (alkyl)carbonyl that is optionally substitutedwith 1-3 of halo, hydroxy, or combinations thereof. In other examples,R₃ is (methyl)carbonyl, (ethyl)carbonyl, (propyl)carbonyl, or(butyl)carbonyl, each of which is optionally substituted with 1-3 ofhalo, hydroxy, or combinations thereof.

In several embodiments, R₃ is an optionally substituted(cycloaliphatic)carbonyl or an optionally substituted(heterocycloaliphatic)carbonyl. In several examples, R₃ is an optionallysubstituted (C₃₋₇ cycloaliphatic)carbonyl. For example, R₃ is a(cyclopropyl)carbonyl, (cyclobutyl)carbonyl, (cyclopentyl)carbonyl,(cyclohexyl)carbonyl, or (cycloheptyl)carbonyl, each of which isoptionally substituted with aliphatic, halo, hydroxy, nitro, cyano, orcombinations thereof. In several alternative examples, R₃ is anoptionally substituted (heterocycloaliphatic)carbonyl. For example, R₃is an optionally substituted (heterocycloaliphatic)carbonyl having 1-3heteroatoms independently selected from N, O, and S. In other examples,R₃ is an optionally substituted (heterocycloaliphatic)carbonyl having1-3 heteroatoms independently selected from N and O. In still otherexamples, R₃ is an optionally substituted 4-7 membered monocyclic(heterocycloaliphatic)carbonyl having 1-3 heteroatoms independentlyselected from N and O. Alternatively, R₃ is (piperidine-1-yl,)carbonyl,(pyrrolidine-1-yl)carbonyl, or (morpholine-4-yl)carbonyl,(piperazine-1-yl)carbonyl, each of which is optionally substituted with1-3 of halo, hydroxy, cyano, nitro, or aliphatic.

In still other instances, R₃ is optionally substituted (aliphatic)amidosuch as (aliphatic(amino(carbonyl)) that is attached to the 2 or 3position on the indole ring of formula Ia. In some embodiments, R₃ is anoptionally substituted (alkyl(amino))carbonyl that is attached to the 2or 3 position on the indole ring of formula Ia. In other embodiments, R₃is an optionally substituted straight or branched(aliphatic(amino))carbonyl that is attached to the 2 or 3 position onthe indole ring of formula Ia. In several examples, R₃ is(N,N-dimethyl(amino))carbonyl, (methyl(amino))carbonyl,(ethyl(amino))carbonyl, (propyl(amino))carbonyl,(prop-2-yl(amino))carbonyl, (dimethyl(but-2-yl(amino)))carbonyl,(tertbutyl(amino))carbonyl, (butyl(amino))carbonyl, each of which isoptionally substituted with 1-3 of halo, hydroxy, cycloaliphatic,heterocycloaliphatic, aryl, heteroaryl, or combinations thereof.

In other embodiments, R₃ is an optionally substituted (alkoxy)carbonyl.For example, R₃ is (methoxy)carbonyl, (ethoxy)carbonyl,(propoxy)carbonyl, or (butoxy)carbonyl, each of which is optionallysubstituted with 1-3 of halo, hydroxy, or combinations thereof. Inseveral instances, R₃ is an optionally substituted straight or branchedC₁₋₆ aliphatic. For example, R₃ is an optionally substituted straight orbranched C₁₋₆ alkyl. In other examples, R₃ is independently anoptionally substituted methyl, ethyl, propyl, butyl, isopropyl, ortertbutyl, each of which is optionally substituted with 1-3 of halo,hydroxy, cyano, nitro, or combination thereof. In other embodiments, R₃is an optionally substituted C₃₋₆ cycloaliphatic. Exemplary embodimentsinclude cyclopropyl, 1-methyl-cycloprop-1-yl, etc. In other examples, pis 2 and the two R₃ substituents are attached to the indole of formulaIa at the 2,4- or 2,6- or 2,7-positions. Exemplary embodiments include6-F, 3-(optionally substituted C₁₋₆ aliphatic or C₃₋₆ cycloaliphatic);7-F-2-(-(optionally substituted C₁₋₆ aliphatic or C₃₋₆ cycloaliphatic)),4F-2-(optionally substituted C₁₋₆ aliphatic or C₃₋₆ cycloaliphatic);7-CN-2-(optionally substituted C₁₋₆ aliphatic or C₃₋₆ cycloaliphatic);7-Me-2-(optionally substituted C₁₋₆ aliphatic or C₃₋₆ cycloaliphatic)and 7-OMe-2-(optionally substituted C₁₋₆ aliphatic or C₃₋₆cycloaliphatic).

In several embodiments, R₃ is hydrogen. In several instances, R₃ is anoptionally substituted straight or branched C₁₋₆ aliphatic. In otherembodiments, R₃ is an optionally substituted C₃₋₆ cycloaliphatic.

In several embodiments, R₃ is one selected from: —H, —CH₃, —CH₂OH,—CH₂CH₃, —CH₂CH₂OH, —CH₂CH₂CH₃, —NH₂, halo, —OCH₃, —CN, —CF₃,—C(O)OCH₂CH₃, —S(O)₂CH₃, —CH₂NH₂, —C(O)NH₂,

In another embodiment, two adjacent R₃ groups form

In several embodiments, R′₃ is independently —Z^(C)R₆, where each Z^(C)is independently a bond or an optionally substituted branched orstraight C₁₋₆ aliphatic chain wherein up to two carbon units of Z^(C)are optionally and independently replaced by —CO—, —CS—, —CONR^(C)—,—CONR^(C)NR^(C)—, —CO₂—, —OCO—, —NR^(C)CO₂—, —O—, —NR^(C)CONR^(C)—,—OCONR^(C)—, —NR^(C)NR^(C)—, NR^(C)CO—, —S—, —SO—, —SO₂—, —NR^(C)—,—SO₂NR^(C)—, —NR^(C)SO₂—, or —NR^(C)SO₂NR^(C)—. Each R₆ is independentlyR^(C), halo, —OH, —NH₂, —NO₂, —CN, or —OCF₃. Each R^(C) is independentlyhydrogen, an optionally substituted aliphatic, an optionally substitutedcycloaliphatic, an optionally substituted heterocycloaliphatic, or anoptionally substituted heteroaryl. In one embodiment, each R^(C) ishydrogen, C₁₋₆ aliphatic, or C₃₋₆ cycloaliphatic, wherein either of thealiphatic or cycloaliphatic is optionally substituted with up to 4-OHsubstituents. In another embodiment, R^(C) is hydrogen, or C₁₋₆ alkyloptionally substituted with up to 4-OH substituents.

For example, in many embodiments, R′₃ is independently —Z^(C)R₆, whereeach Z^(C) is independently a bond or an optionally substituted branchedor straight C₁₋₆ aliphatic chain wherein up to two carbon units of Z^(C)are optionally and independently replaced by —C(O)—, —C(O)NR^(C)—,—C(O)O—, —NR^(C)C(O)O—, —O—, —NR^(C)S(O)₂—, or —NR^(C)—. Each R₆ isindependently R^(C),

—OH, or —NH₂. Each R^(C) is independently hydrogen, an optionallysubstituted cycloaliphatic, an optionally substitutedheterocycloaliphatic, or an optionally substituted heteroaryl. In oneembodiment, each R^(C) is hydrogen, C₁₋₆ aliphatic, or C₃₋₆cycloaliphatic, wherein either of the aliphatic or cycloaliphatic isoptionally substituted with up to 4-OH substituents. In anotherembodiment, R^(C) is hydrogen, or C₁₋₆ alkyl optionally substituted withup to 4-OH substituents.

In other embodiments, R′₃ is hydrogen or

wherein R₃₁ is H or a C₁₋₂ aliphatic that is optionally substituted with1-3 of halo, —OH, or combinations thereof. R₃₂ is -L-R₃₃, wherein L is abond, —CH₂—, —CH₂O—, —CH₂NHS(O)₂—, —CH₂C(O)—, —CH₂NHC(O)—, or —CH₂NH—;and R₃₃ is hydrogen, or C₁₋₂ aliphatic, cycloaliphatic,heterocycloaliphatic, or heteroaryl, each of which is optionallysubstituted with 1 of —OH,

—NH₂, or —CN. For example, in one embodiment, R₃₁ is hydrogen and R₃₂ isC₁₋₂ aliphatic optionally substituted with —OH, —NH₂, or —CN.

In several embodiments, R′₃ is independently selected from one of thefollowing: —H, —CH₃, —CH₂CH₃, —C(O)CH₃, —CH₂CH₂OH, —C(O)OCH₃,

5. n Term

n is 1-3.

In several embodiments, n is 1. In other embodiments, n is 2. In stillother embodiments, n is 3.

C. Exemplary Compounds of the Present Invention

Exemplary compounds of the present invention include, but are notlimited to, those illustrated in Table 1 below.

TABLE 1 Exemplary compounds of the present invention.

1

2

3

4

5

6

7

8

9

10

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305

306

III. Subgeneric Compounds of the Present Invention

Another aspect of the present invention provides a compound that isuseful for modulating ABC transporter activity. The compound has formulaIc:

or a pharmaceutically acceptable salt thereof.

R₁, R₂, and ring A are defined above in formula I, and ring B, R₃ and pare defined in formula Ia. Furthermore, when ring A is unsubstitutedcyclopentyl, n is 1, R₂ is 4-chloro, and R₁ is hydrogen, then ring B isnot 2-(tertbutyl)indol-5-yl, or(2,6-dichlorophenyl(carbonyl))-3-methyl-1H-indol-5-yl; and when ring Ais unsubstituted cyclopentyl, n is 0, and R₁ is hydrogen, then ring B isnot

Another aspect of the present invention provides a compound that isuseful for modulating ABC transporter activity. The compound has formulaId.

or a pharmaceutically acceptable salt thereof.

R₁, R₂, and ring A are defined above in formula I, and ring B, R₃ and pare defined in formula Ia.

However, when R₁ is H, n is 0, ring A is an unsubstituted cyclopentyl,and ring B is an indole-5-yl substituted with 1-2 of R₃, then each R₃ isindependently —Z^(G)R₁₂, where each Z^(G) is independently a bond or anunsubstituted branched or straight C₁₋₆ aliphatic chain wherein up totwo carbon units of Z^(G) are optionally and independently replaced by—CS—, —CONR^(G)NR^(G)—, —CO₂—, —OCO—, —NR^(G)CO₂—, —O—,—NR^(G)CONR^(G)—, —OCONR^(G)—, —NR^(G)NR^(G)—, —S-, —SO—, —SO₂—,—NR^(G),

—SO₂NR^(G)—, —NR^(G)SO₂—, or —NR^(G)SO₂NR^(C)—, each R₁₂ isindependently R^(G), halo, —OH, —NH₂, —NO₂, —CN, or —OCF₃, and eachR^(G) is independently hydrogen, an unsubstituted aliphatic, anoptionally substituted cycloaliphatic, an optionally substitutedheterocycloaliphatic, an unsubstituted aryl, or an optionallysubstituted heteroaryl; or any two adjacent R₃ groups together with theatoms to which they are attached form an optionally substitutedheterocycle. Furthermore, when R₁ is H, n is 1, R₂ is 4-chloro, ring Ais an unsubstituted cyclopentyl, and ring B is an indole-5-ylsubstituted with 1-2 of R₃, then each R₃ is independently —Z^(H)R₂₂,where each Z^(H) is independently a bond or an unsubstituted branched orstraight C₁₋₃ aliphatic chain wherein up to two carbon units of Z^(H)are optionally and independently replaced by —CS—, —CONR^(H)NR^(H),—CO₂—, —OCO—,—NR^(H)CO₂—, —O—, —NR^(H)CONR^(H)—, —OCONR^(H)—, —NR^(H)NR^(H)—, —S—,—SO—, —SO₂—, —NR^(H)—, —SO₂NR^(H)—, —NR^(H)SO₂—, or —NR^(H)SO₂NR^(H)—,each R₂₂ is independently R^(H), halo, —OH, —NH₂, —NO₂, —CN, or —OCF₃,and each R^(H) is independently hydrogen, a substituted C₄ alkyl, anoptionally substituted C₂₋₆ alkenyl, an optionally substituted C₂₋₆alkynyl, an optionally substituted C₄ alkenyl, an optionally substitutedC₄ alkynyl, an optionally substituted cycloaliphatic, an optionallysubstituted heterocycloaliphatic, an optionally substituted heteroaryl,an unsubstituted phenyl, or a mono-substituted phenyl, or any twoadjacent R₃ groups together with the atoms to which they are attachedform an optionally substituted heterocycle.

Another aspect of the present invention provides a compound that isuseful for modulating ABC transporter activity. The compound has formulaII:

or a pharmaceutically acceptable salt thereof.

R₁, R₂, and ring A are defined above in formula I; R₃, R′₃, and p aredefined above in formula Ia; and Z₁, Z₂, Z₃, Z4, and Z₅ are definedabove in formula Ib.

Another aspect of the present invention provides a compound that isuseful for modulating ABC transporter activity. The compound has formulaIIa:

or a pharmaceutically acceptable salt thereof.

R₁, R₂, and ring A are defined above in formula I; R₃, R′₃, and p aredefined above in formula Ia; and Z₁, Z₂, Z₃, Z4, and Z₅ are definedabove in formula Ib.

Another aspect of the present invention provides a compound that isuseful for modulating ABC transporter activity. The compound has formulaIIb:

or a pharmaceutically acceptable salt thereof.

R₁, R₂, and ring A, are defined above in formula I; R₃, R′₃, and p aredefined above in formula Ia; and Z₁, Z₂, Z₃, Z4, and Z₅ are definedabove in formula Ib.

Another aspect of the present invention provides a compound that isuseful for modulating ABC transporter activity. The compound has formulaIIc:

or a pharmaceutically acceptable salt thereof.

R₁, R₂ and n are defined above in formula I; and R₃, R′₃, and p aredefined in formula Ia.

Another aspect of the present invention provides a compound that isuseful for modulating ABC transporter activity. The compound has formulaIId:

or a pharmaceutically acceptable salt thereof.

Both R₂ groups, together with the atoms to which they are attached forma group selected from:

R′₃ is independently selected from one of the following:

—H, —CH₃, —CH₂CH₃, —C(O)CH₃, —CH₂CH₂OH, —C(O)—OCH₃,

and each R₃ is independently selected from —H, —CH₃, —CH₂OH, —CH₂CH₃,—CH₂CH₂OH, —CH₂CH₂CH₃, —NH₂, halo, —OCH₃, —CN, —CF₃,—C(O)OCH₂CH₃, —S(O)₂CH₃, —CH₂NH₂, —C(O)NH₂,

IV. Generic Synthetic Schemes

The compounds of formulae (I, Ic, Id, II, IIa, IIb, IIc, and IId) may bereadily synthesized from commercially available or known startingmaterials by known methods. Exemplary synthetic routes to producecompounds of formulae (I, Ic, Id, II, IIa, IIb, IIc, and IId) areprovided below in Schemes 1-22 below.

Preparation of the compounds of the invention is achieved by thecoupling of a ring B amine with a ring A carboxylic acid as illustratedin Scheme 1.

Referring to Scheme 1, the acid 1a may be converted to the correspondingacid chloride 1b using thionyl chloride in the presence of a catalysticamount of dimethylformamide. Reaction of the acid chloride with theamine

provides compounds of the invention I. Alternatively, the acid 1a may bedirectly coupled to the amine using known coupling reagents such as, forexample, HATU in the presence of triethylamine.

Preparation of the acids 1a may be achieved as illustrated in Scheme 2.

Referring to Scheme 2, the nitrile 2a reacts with a suitablebromochloroalkane in the presence of sodium hydroxide and a phasetransfer catalyst such as butyltriethylammonium chloride to provide theintermediate 2b. Hydrolysis of the nitrile of 2b provides the acid 1a.In some instances, isolation of the intermediate 2b is unnecessary.

The phenylacetonitriles 2a are commercially available or may be preparedas illustrated in Scheme 3.

-   -   a) Pd(PPh₃)₄, CO, MeOH; b) LiAlH₄, THF; c) SOCl₂; d) NaCN

Referring to Scheme 3, reaction of an aryl bromide 3a with carbonmonoxide in the presence of methanol andtetrakis(triphenylphosphine)palladium (0) provides the ester 3b.Reduction of 3b with lithium aluminum hydride provides the alcohol 3cwhich is converted to the halide 3d with thionyl chloride. Reaction of3d with sodium cyanide provides the nitrile 2a.

Other methods of producing the nitrile 2a are illustrated in schemes 4and 5 below.

Preparation of

components is illustrated in the schemes that follow. A number ofmethods for preparing ring B compounds wherein ring B is an indole havebeen reported. See for example Angew. Chem. 2005, 44, 606; J. Am. Chem.Soc. 2005, 127, 5342,); J. Comb. Chem. 2005, 7, 130; Tetrahedron 2006,62, 3439; J. Chem. Soc. Perkin Trans. 1, 2000, 1045.

One method for preparing

is illustrated in Scheme 6.

Referring to Scheme 6, a nitroaniline 6a is converted to the hydrazine6b using nitrous acid in the presence of HCl and stannous chloride.Reaction of 6b with an aldehyde or ketone CH₃C(O)R₃ provides thehydrazone 6c which on treatment with phosphoric acid in toluene leads toa mixture of nitro indoles 6d and 6e. Catalytic hydrogenation in thepresence of palladium on carbon provides a mixture of the amino indoles6f and 6g which may be separated using know methods such as, forexample, chromatography.

An alternative method is illustrated in scheme 7.

In the schemes above, the radical R employed therein is a substituent,e.g., RW as defined hereinabove. One of skill in the art will readilyappreciate that synthetic routes suitable for various substituents ofthe present invention are such that the reaction conditions and stepsemployed do not modify the intended substituents.

V. Formulations, Administrations, and Uses

Accordingly, in another aspect of the present invention,pharmaceutically acceptable compositions are provided, wherein thesecompositions comprise any of the compounds as described herein, andoptionally comprise a pharmaceutically acceptable carrier, adjuvant orvehicle. In certain embodiments, these compositions optionally furthercomprise one or more additional therapeutic agents.

It will also be appreciated that certain of the compounds of presentinvention can exist in free form for treatment, or where appropriate, asa pharmaceutically acceptable derivative or a prodrug thereof. Accordingto the present invention, a pharmaceutically acceptable derivative or aprodrug includes, but is not limited to, pharmaceutically acceptablesalts, esters, salts of such esters, or any other adduct or derivativewhich upon administration to a patient in need is capable of providing,directly or indirectly, a compound as otherwise described herein, or ametabolite or residue thereof.

As used herein, the term “pharmaceutically acceptable salt” refers tothose salts which are, within the scope of sound medical judgment,suitable for use in contact with the tissues of humans and lower animalswithout undue toxicity, irritation, allergic response and the like, andare commensurate with a reasonable benefit/risk ratio. A“pharmaceutically acceptable salt” means any salt or salt of an ester ofa compound of this invention that, upon administration to a recipient,is capable of providing, either directly or indirectly, a compound ofthis invention or an inhibitorily active metabolite or residue thereof.

Pharmaceutically acceptable salts are well known in the art. Forexample, S. M. Berge, et al. describes pharmaceutically acceptable saltsin detail in J. Pharmaceutical Sciences, 1977, 66, 1-19, incorporatedherein by reference. Pharmaceutically acceptable salts of the compoundsof this invention include those derived from suitable inorganic andorganic acids and bases. Examples of pharmaceutically acceptable,nontoxic acid addition salts are salts of an amino group formed withinorganic acids such as hydrochloric acid, hydrobromic acid, phosphoricacid, sulfuric acid and perchloric acid or with organic acids such asacetic acid, oxalic acid, maleic acid, tartaric acid, citric acid,succinic acid or malonic acid or by using other methods used in the artsuch as ion exchange. Other pharmaceutically acceptable salts includeadipate, alginate, ascorbate, aspartate, benzenesulfonate, benzoate,bisulfate, borate, butyrate, camphorate, camphorsulfonate, citrate,cyclopentanepropionate, digluconate, dodecylsulfate, ethanesulfonate,formate, fumarate, glucoheptonate, glycerophosphate, gluconate,hemisulfate, heptanoate, hexanoate, hydroiodide,2-hydroxy-ethanesulfonate, lactobionate, lactate, laurate, laurylsulfate, malate, maleate, malonate, methanesulfonate,2-naphthalenesulfonate, nicotinate, nitrate, oleate, oxalate, palmitate,pamoate, pectinate, persulfate, 3-phenylpropionate, phosphate, picrate,pivalate, propionate, stearate, succinate, sulfate, tartrate,thiocyanate, p-toluenesulfonate, undecanoate, valerate salts, and thelike. Salts derived from appropriate bases include alkali metal,alkaline earth metal, ammonium and N⁺(C₁₋₄alkyl)₄ salts. This inventionalso envisions the quaternization of any basic nitrogen-containinggroups of the compounds disclosed herein. Water or oil-soluble ordispersible products may be obtained by such quaternization.Representative alkali or alkaline earth metal salts include sodium,lithium, potassium, calcium, magnesium, and the like. Furtherpharmaceutically acceptable salts include, when appropriate, nontoxicammonium, quaternary ammonium, and amine cations formed usingcounterions such as halide, hydroxide, carboxylate, sulfate, phosphate,nitrate, loweralkyl sulfonate and aryl sulfonate.

As described above, the pharmaceutically acceptable compositions of thepresent invention additionally comprise a pharmaceutically acceptablecarrier, adjuvant, or vehicle, which, as used herein, includes any andall solvents, diluents, or other liquid vehicle, dispersion orsuspension aids, surface active agents, isotonic agents, thickening oremulsifying agents, preservatives, solid binders, lubricants and thelike, as suited to the particular dosage form desired. Remington'sPharmaceutical Sciences, Sixteenth Edition, E. W. Martin (MackPublishing Co., Easton, Pa., 1980) discloses various carriers used informulating pharmaceutically acceptable compositions and knowntechniques for the preparation thereof. Except insofar as anyconventional carrier medium is incompatible with the compounds of theinvention, such as by producing any undesirable biological effect orotherwise interacting in a deleterious manner with any othercomponent(s) of the pharmaceutically acceptable composition, its use iscontemplated to be within the scope of this invention. Some examples ofmaterials which can serve as pharmaceutically acceptable carriersinclude, but are not limited to, ion exchangers, alumina, aluminumstearate, lecithin, serum proteins, such as human serum albumin, buffersubstances such as phosphates, glycine, sorbic acid, or potassiumsorbate, partial glyceride mixtures of saturated vegetable fatty acids,water, salts or electrolytes, such as protamine sulfate, disodiumhydrogen phosphate, potassium hydrogen phosphate, sodium chloride, zincsalts, colloidal silica, magnesium trisilicate, polyvinyl pyrrolidone,polyacrylates, waxes, polyethylene-polyoxypropylene-block polymers, woolfat, sugars such as lactose, glucose and sucrose; starches such as cornstarch and potato starch; cellulose and its derivatives such as sodiumcarboxymethyl cellulose, ethyl cellulose and cellulose acetate; powderedtragacanth; malt; gelatin; talc; excipients such as cocoa butter andsuppository waxes; oils such as peanut oil, cottonseed oil; saffloweroil; sesame oil; olive oil; corn oil and soybean oil; glycols; such apropylene glycol or polyethylene glycol; esters such as ethyl oleate andethyl laurate; agar; buffering agents such as magnesium hydroxide andaluminum hydroxide; alginic acid; pyrogen-free water; isotonic saline;Ringer's solution; ethyl alcohol, and phosphate buffer solutions, aswell as other non-toxic compatible lubricants such as sodium laurylsulfate and magnesium stearate, as well as coloring agents, releasingagents, coating agents, sweetening, flavoring and perfuming agents,preservatives and antioxidants can also be present in the composition,according to the judgment of the formulator.

In yet another aspect, the present invention provides a method oftreating a condition, disease, or disorder implicated by ABC transporteractivity. In certain embodiments, the present invention provides amethod of treating a condition, disease, or disorder implicated by adeficiency of ABC transporter activity, the method comprisingadministering a composition comprising a compound of formulae (I, Ic,Id, II, IIa, IIb, IIc, and IId) to a subject, preferably a mammal, inneed thereof.

In certain preferred embodiments, the present invention provides amethod of treating Cystic fibrosis, Hereditary emphysema, Hereditaryhemochromatosis, Coagulation-Fibrinolysis deficiencies, such as ProteinC deficiency, Type 1 hereditary angioedema, Lipid processingdeficiencies, such as Familial hypercholesterolemia, Type 1chylomicronemia, Abetalipoproteinemia, Lysosomal storage diseases, suchas I-cell disease/Pseudo-Hurler, Mucopolysaccharidoses,Sandhof/Tay-Sachs, Crigler-Najjar type II,Polyendocrinopathy/Hyperinsulemia, Diabetes mellitus, Laron dwarfism,Myleoperoxidase deficiency, Primary hypoparathyroidism, Melanoma,Glycanosis CDG type 1, Hereditary emphysema, Congenital hyperthyroidism,Osteogenesis imperfecta, Hereditary hypofibrinogenemia, ACT deficiency,Diabetes insipidus (DI), Neurophyseal DI, Neprogenic DI, Charcot-MarieTooth syndrome, Perlizaeus-Merzbacher disease, neurodegenerativediseases such as Alzheimer's disease, Parkinson's disease, Amyotrophiclateral sclerosis, Progressive supranuclear plasy, Pick's disease,several polyglutamine neurological disorders asuch as Huntington,Spinocerebullar ataxia type I, Spinal and bulbar muscular atrophy,Dentatorubal pallidoluysian, and Myotonic dystrophy, as well asSpongiform encephalopathies, such as Hereditary Creutzfeldt-Jakobdisease (due to Prion protein processing defect), Fabry disease,Straussler-Scheinker disease, secretory diarrhea, polycystic kidneydisease, chronic obstructive pulmonary disease (COPD), dry eye disease,and Sjögren's Syndrome, comprising the step of administering to saidmammal an effective amount of a composition comprising a compound offormulae (I, Ic, Id, II, IIa, IIb, IIc, and IId), or a preferredembodiment thereof as set forth above.

According to an alternative preferred embodiment, the present inventionprovides a method of treating cystic fibrosis comprising the step ofadministering to said mammal a composition comprising the step ofadministering to said mammal an effective amount of a compositioncomprising a compound of formulae (I, Ic, Id, II, IIa, IIb, IIc, andIId), or a preferred embodiment thereof as set forth above.

According to the invention an “effective amount” of the compound orpharmaceutically acceptable composition is that amount effective fortreating or lessening the severity of one or more of Cystic fibrosis,Hereditary emphysema, Hereditary hemochromatosis,Coagulation-Fibrinolysis deficiencies, such as Protein C deficiency,Type 1 hereditary angioedema, Lipid processing deficiencies, such asFamilial hypercholesterolemia, Type 1 chylomicronemia,Abetalipoproteinemia, Lysosomal storage diseases, such as I-celldisease/Pseudo-Hurler, Mucopolysaccharidoses, Sandhof/Tay-Sachs,Crigler-Najjar type II, Polyendocrinopathy/Hyperinsulemia, Diabetesmellitus, Laron dwarfism, Myleoperoxidase deficiency, Primaryhypoparathyroidism, Melanoma, Glycanosis CDG type 1, Hereditaryemphysema, Congenital hyperthyroidism, Osteogenesis imperfecta,Hereditary hypofibrinogenemia, ACT deficiency, Diabetes insipidus (DI),Neurophyseal DI, Neprogenic DI, Charcot-Marie Tooth syndrome,Perlizaeus-Merzbacher disease, neurodegenerative diseases such asAlzheimer's disease, Parkinson's disease, Amyotrophic lateral sclerosis,Progressive supranuclear plasy, Pick's disease, several polyglutamineneurological disorders asuch as Huntington, Spinocerebullar ataxia typeI, Spinal and bulbar muscular atrophy, Dentatorubal pallidoluysian, andMyotonic dystrophy, as well as Spongiform encephalopathies, such asHereditary Creutzfeldt-Jakob disease, Fabry disease,Straussler-Scheinker disease, secretory diarrhea, polycystic kidneydisease, chronic obstructive pulmonary disease (COPD), dry eye disease,and Sjögren's Syndrome.

The compounds and compositions, according to the method of the presentinvention, may be administered using any amount and any route ofadministration effective for treating or lessening the severity of oneor more of Cystic fibrosis, Hereditary emphysema, Hereditaryhemochromatosis, Coagulation-Fibrinolysis deficiencies, such as ProteinC deficiency, Type 1 hereditary angioedema, Lipid processingdeficiencies, such as Familial hypercholesterolemia, Type 1chylomicronemia, Abetalipoproteinemia, Lysosomal storage diseases, suchas I-cell disease/Pseudo-Hurler, Mucopolysaccharidoses,Sandhof/Tay-Sachs, Crigler-Najjar type II,Polyendocrinopathy/Hyperinsulemia, Diabetes mellitus, Laron dwarfism,Myleoperoxidase deficiency, Primary hypoparathyroidism, Melanoma,Glycanosis CDG type 1, Hereditary emphysema, Congenital hyperthyroidism,Osteogenesis imperfecta, Hereditary hypofibrinogenemia, ACT deficiency,Diabetes insipidus (DI), Neurophyseal DI, Neprogenic DI, Charcot-MarieTooth syndrome, Perlizaeus-Merzbacher disease, neurodegenerativediseases such as Alzheimer's disease, Parkinson's disease, Amyotrophiclateral sclerosis, Progressive supranuclear plasy, Pick's disease,several polyglutamine neurological disorders asuch as Huntington,Spinocerebullar ataxia type I, Spinal and bulbar muscular atrophy,Dentatorubal pallidoluysian, and Myotonic dystrophy, as well asSpongiform encephalopathies, such as Hereditary Creutzfeldt-Jakobdisease, Fabry disease, Straussler-Scheinker disease, secretorydiarrhea, polycystic kidney disease, chronic obstructive pulmonarydisease (COPD), dry eye disease, and Sjögren's Syndrome.

The exact amount required will vary from subject to subject, dependingon the species, age, and general condition of the subject, the severityof the infection, the particular agent, its mode of administration, andthe like. The compounds of the invention are preferably formulated indosage unit form for ease of administration and uniformity of dosage.The expression “dosage unit form” as used herein refers to a physicallydiscrete unit of agent appropriate for the patient to be treated. Itwill be understood, however, that the total daily usage of the compoundsand compositions of the present invention will be decided by theattending physician within the scope of sound medical judgment. Thespecific effective dose level for any particular patient or organismwill depend upon a variety of factors including the disorder beingtreated and the severity of the disorder; the activity of the specificcompound employed; the specific composition employed; the age, bodyweight, general health, sex and diet of the patient; the time ofadministration, route of administration, and rate of excretion of thespecific compound employed; the duration of the treatment; drugs used incombination or coincidental with the specific compound employed, andlike factors well known in the medical arts. The term “patient”, as usedherein, means an animal, preferably a mammal, and most preferably ahuman.

The pharmaceutically acceptable compositions of this invention can beadministered to humans and other animals orally, rectally, parenterally,intracisternally, intravaginally, intraperitoneally, topically (as bypowders, ointments, or drops), bucally, as an oral or nasal spray, orthe like, depending on the severity of the infection being treated. Incertain embodiments, the compounds of the invention may be administeredorally or parenterally at dosage levels of about 0.01 mg/kg to about 50mg/kg and preferably from about 1 mg/kg to about 25 mg/kg, of subjectbody weight per day, one or more times a day, to obtain the desiredtherapeutic effect.

Liquid dosage forms for oral administration include, but are not limitedto, pharmaceutically acceptable emulsions, microemulsions, solutions,suspensions, syrups and elixirs. In addition to the active compounds,the liquid dosage forms may contain inert diluents commonly used in theart such as, for example, water or other solvents, solubilizing agentsand emulsifiers such as ethyl alcohol, isopropyl alcohol, ethylcarbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propyleneglycol, 1,3-butylene glycol, dimethylformamide, oils (in particular,cottonseed, groundnut, corn, germ, olive, castor, and sesame oils),glycerol, tetrahydrofurfuryl alcohol, polyethylene glycols and fattyacid esters of sorbitan, and mixtures thereof. Besides inert diluents,the oral compositions can also include adjuvants such as wetting agents,emulsifying and suspending agents, sweetening, flavoring, and perfumingagents.

Injectable preparations, for example, sterile injectable aqueous oroleaginous suspensions may be formulated according to the known artusing suitable dispersing or wetting agents and suspending agents. Thesterile injectable preparation may also be a sterile injectablesolution, suspension or emulsion in a nontoxic parenterally acceptablediluent or solvent, for example, as a solution in 1,3-butanediol. Amongthe acceptable vehicles and solvents that may be employed are water,Ringer's solution, U.S.P. and isotonic sodium chloride solution. Inaddition, sterile, fixed oils are conventionally employed as a solventor suspending medium. For this purpose any bland fixed oil can beemployed including synthetic mono- or diglycerides. In addition, fattyacids such as oleic acid are used in the preparation of injectables.

The injectable formulations can be sterilized, for example, byfiltration through a bacterial-retaining filter, or by incorporatingsterilizing agents in the form of sterile solid compositions which canbe dissolved or dispersed in sterile water or other sterile injectablemedium prior to use.

In order to prolong the effect of a compound of the present invention,it is often desirable to slow the absorption of the compound fromsubcutaneous or intramuscular injection. This may be accomplished by theuse of a liquid suspension of crystalline or amorphous material withpoor water solubility. The rate of absorption of the compound thendepends upon its rate of dissolution that, in turn, may depend uponcrystal size and crystalline form. Alternatively, delayed absorption ofa parenterally administered compound form is accomplished by dissolvingor suspending the compound in an oil vehicle. Injectable depot forms aremade by forming microencapsule matrices of the compound in biodegradablepolymers such as polylactide-polyglycolide. Depending upon the ratio ofcompound to polymer and the nature of the particular polymer employed,the rate of compound release can be controlled. Examples of otherbiodegradable polymers include poly(orthoesters) and poly(anhydrides).Depot injectable formulations are also prepared by entrapping thecompound in liposomes or microemulsions that are compatible with bodytissues.

Compositions for rectal or vaginal administration are preferablysuppositories which can be prepared by mixing the compounds of thisinvention with suitable non-irritating excipients or carriers such ascocoa butter, polyethylene glycol or a suppository wax which are solidat ambient temperature but liquid at body temperature and therefore meltin the rectum or vaginal cavity and release the active compound.

Solid dosage forms for oral administration include capsules, tablets,pills, powders, and granules. In such solid dosage forms, the activecompound is mixed with at least one inert, pharmaceutically acceptableexcipient or carrier such as sodium citrate or dicalcium phosphateand/or a) fillers or extenders such as starches, lactose, sucrose,glucose, mannitol, and silicic acid, b) binders such as, for example,carboxymethylcellulose, alginates, gelatin, polyvinylpyrrolidinone,sucrose, and acacia, c) humectants such as glycerol, d) disintegratingagents such as agar—agar, calcium carbonate, potato or tapioca starch,alginic acid, certain silicates, and sodium carbonate, e) solutionretarding agents such as paraffin, f) absorption accelerators such asquaternary ammonium compounds, g) wetting agents such as, for example,cetyl alcohol and glycerol monostearate, h) absorbents such as kaolinand bentonite clay, and i) lubricants such as talc, calcium stearate,magnesium stearate, solid polyethylene glycols, sodium lauryl sulfate,and mixtures thereof. In the case of capsules, tablets and pills, thedosage form may also comprise buffering agents.

Solid compositions of a similar type may also be employed as fillers insoft and hard-filled gelatin capsules using such excipients as lactoseor milk sugar as well as high molecular weight polyethylene glycols andthe like. The solid dosage forms of tablets, dragees, capsules, pills,and granules can be prepared with coatings and shells such as entericcoatings and other coatings well known in the pharmaceutical formulatingart. They may optionally contain opacifying agents and can also be of acomposition that they release the active ingredient(s) only, orpreferentially, in a certain part of the intestinal tract, optionally,in a delayed manner. Examples of embedding compositions that can be usedinclude polymeric substances and waxes. Solid compositions of a similartype may also be employed as fillers in soft and hard-filled gelatincapsules using such excipients as lactose or milk sugar as well as highmolecular weight polethylene glycols and the like.

The active compounds can also be in microencapsulated form with one ormore excipients as noted above. The solid dosage forms of tablets,dragees, capsules, pills, and granules can be prepared with coatings andshells such as enteric coatings, release controlling coatings and othercoatings well known in the pharmaceutical formulating art. In such soliddosage forms the active compound may be admixed with at least one inertdiluent such as sucrose, lactose or starch. Such dosage forms may alsocomprise, as is normal practice, additional substances other than inertdiluents, e.g., tableting lubricants and other tableting aids such amagnesium stearate and microcrystalline cellulose. In the case ofcapsules, tablets and pills, the dosage forms may also comprisebuffering agents. They may optionally contain opacifying agents and canalso be of a composition that they release the active ingredient(s)only, or preferentially, in a certain part of the intestinal tract,optionally, in a delayed manner. Examples of embedding compositions thatcan be used include polymeric substances and waxes.

Dosage forms for topical or transdermal administration of a compound ofthis invention include ointments, pastes, creams, lotions, gels,powders, solutions, sprays, inhalants or patches. The active componentis admixed under sterile conditions with a pharmaceutically acceptablecarrier and any needed preservatives or buffers as may be required.Ophthalmic formulation, eardrops, and eye drops are also contemplated asbeing within the scope of this invention. Additionally, the presentinvention contemplates the use of transdermal patches, which have theadded advantage of providing controlled delivery of a compound to thebody. Such dosage forms are prepared by dissolving or dispensing thecompound in the proper medium. Absorption enhancers can also be used toincrease the flux of the compound across the skin. The rate can becontrolled by either providing a rate controlling membrane or bydispersing the compound in a polymer matrix or gel.

As described generally above, the compounds of the invention are usefulas modulators of ABC transporters. Thus, without wishing to be bound byany particular theory, the compounds and compositions are particularlyuseful for treating or lessening the severity of a disease, condition,or disorder where hyperactivity or inactivity of ABC transporters isimplicated in the disease, condition, or disorder. When hyperactivity orinactivity of an ABC transporter is implicated in a particular disease,condition, or disorder, the disease, condition, or disorder may also bereferred to as a “ABC transporter-mediated disease, condition ordisorder”. Accordingly, in another aspect, the present inventionprovides a method for treating or lessening the severity of a disease,condition, or disorder where hyperactivity or inactivity of an ABCtransporter is implicated in the disease state.

The activity of a compound utilized in this invention as a modulator ofan ABC transporter may be assayed according to methods describedgenerally in the art and in the Examples herein.

It will also be appreciated that the compounds and pharmaceuticallyacceptable compositions of the present invention can be employed incombination therapies, that is, the compounds and pharmaceuticallyacceptable compositions can be administered concurrently with, prior to,or subsequent to, one or more other desired therapeutics or medicalprocedures. The particular combination of therapies (therapeutics orprocedures) to employ in a combination regimen will take into accountcompatibility of the desired therapeutics and/or procedures and thedesired therapeutic effect to be achieved. It will also be appreciatedthat the therapies employed may achieve a desired effect for the samedisorder (for example, an inventive compound may be administeredconcurrently with another agent used to treat the same disorder), orthey may achieve different effects (e.g., control of any adverseeffects). As used herein, additional therapeutic agents that arenormally administered to treat or prevent a particular disease, orcondition, are known as “appropriate for the disease, or condition,being treated”.

The amount of additional therapeutic agent present in the compositionsof this invention will be no more than the amount that would normally beadministered in a composition comprising that therapeutic agent as theonly active agent. Preferably the amount of additional therapeutic agentin the presently disclosed compositions will range from about 50% to100% of the amount normally present in a composition comprising thatagent as the only therapeutically active agent.

The compounds of this invention or pharmaceutically acceptablecompositions thereof may also be incorporated into compositions forcoating an implantable medical device, such as prostheses, artificialvalves, vascular grafts, stents and catheters. Accordingly, the presentinvention, in another aspect, includes a composition for coating animplantable device comprising a compound of the present invention asdescribed generally above, and in classes and subclasses herein, and acarrier suitable for coating said implantable device. In still anotheraspect, the present invention includes an implantable device coated witha composition comprising a compound of the present invention asdescribed generally above, and in classes and subclasses herein, and acarrier suitable for coating said implantable device. Suitable coatingsand the general preparation of coated implantable devices are describedin U.S. Pat. Nos. 6,099,562; 5,886,026; and 5,304,121. The coatings aretypically biocompatible polymeric materials such as a hydrogel polymer,polymethyldisiloxane, polycaprolactone, polyethylene glycol, polylacticacid, ethylene vinyl acetate, and mixtures thereof. The coatings mayoptionally be further covered by a suitable topcoat of fluorosilicone,polysaccarides, polyethylene glycol, phospholipids or combinationsthereof to impart controlled release characteristics in the composition.

Another aspect of the invention relates to modulating ABC transporteractivity in a biological sample or a patient (e.g., in vitro or invivo), which method comprises administering to the patient, orcontacting said biological sample with a compound of formula I or acomposition comprising said compound. The term “biological sample”, asused herein, includes, without limitation, cell cultures or extractsthereof, biopsied material obtained from a mammal or extracts thereof;and blood, saliva, urine, feces, semen, tears, or other body fluids orextracts thereof.

Modulation of ABC transporter activity in a biological sample is usefulfor a variety of purposes that are known to one of skill in the art.Examples of such purposes include, but are not limited to, the study ofABC transporters in biological and pathological phenomena; and thecomparative evaluation of new modulators of ABC transporters.

In yet another embodiment, a method of modulating activity of an anionchannel in vitro or in vivo, is provided comprising the step ofcontacting said channel with a compound of formulae (I, Ic, Id, II, IIa,IIb, IIc, and IId). In preferred embodiments, the anion channel is achloride channel or a bicarbonate channel. In other preferredembodiments, the anion channel is a chloride channel.

According to an alternative embodiment, the present invention provides amethod of increasing the number of functional ABC transporters in amembrane of a cell, comprising the step of contacting said cell with acompound of formulae (I, Ic, Id, II, IIa, IIb, IIc, and IId). The term“functional ABC transporter” as used herein means an ABC transporterthat is capable of transport activity. In preferred embodiments, saidfunctional ABC transporter is CFTR.

According to another preferred embodiment, the activity of the ABCtransporter is measured by measuring the transmembrane voltagepotential. Means for measuring the voltage potential across a membranein the biological sample may employ any of the known methods in the art,such as optical membrane potential assay or other electrophysiologicalmethods.

The optical membrane potential assay utilizes voltage-sensitive FRETsensors described by Gonzalez and Tsien (See, Gonzalez, J. E. and R. Y.Tsien (1995) “Voltage sensing by fluorescence resonance energy transferin single cells” Biophys J 69(4): 1272-80, and Gonzalez, J. E. and R. Y.Tsien (1997) “Improved indicators of cell membrane potential that usefluorescence resonance energy transfer” Chem Biol 4(4): 269-77) incombination with instrumentation for measuring fluorescence changes suchas the Voltage/Ion Probe Reader (VIPR) (See, Gonzalez, J. E., K. Oades,et al. (1999) “Cell-based assays and instrumentation for screeningion-channel targets” Drug Discov Today 4(9): 431-439).

These voltage sensitive assays are based on the change in fluorescenceresonant energy transfer (FRET) between the membrane-soluble,voltage-sensitive dye, DiSBAC₂(3), and a fluorescent phospholipid,CC2-DMPE, which is attached to the outer leaflet of the plasma membraneand acts as a FRET donor. Changes in membrane potential (V_(m)) causethe negatively charged DiSBAC₂(3) to redistribute across the plasmamembrane and the amount of energy transfer from CC2-DMPE changesaccordingly. The changes in fluorescence emission can be monitored usingVIPR™ II, which is an integrated liquid handler and fluorescent detectordesigned to conduct cell-based screens in 96- or 384-well microtiterplates.

In another aspect the present invention provides a kit for use inmeasuring the activity of a ABC transporter or a fragment thereof in abiological sample in vitro or in vivo comprising (i) a compositioncomprising a compound of formulae (I, Ic, Id, II, IIa, IIb, IIc, andIId) or any of the above embodiments; and (ii) instructions for a.)contacting the composition with the biological sample and b.) measuringactivity of said ABC transporter or a fragment thereof. In oneembodiment, the kit further comprises instructions for a.) contacting anadditional composition with the biological sample; b.) measuring theactivity of said ABC transporter or a fragment thereof in the presenceof said additional compound, and c.) comparing the activity of the ABCtransporter in the presence of the additional compound with the densityof the ABC transporter in the presence of a composition of formulae (I,Ic, Id, II, IIa, IIb, IIc, and IId). In preferred embodiments, the kitis used to measure the density of CFTR.

In order that the invention described herein may be more fullyunderstood, the following examples are set forth. It should beunderstood that these examples are for illustrative purposes only andare not to be construed as limiting this invention in any manner.

VI. Preparations and Examples

General Procedure I: Carboxylic Acid Building Block

Benzyltriethylammonium chloride (0.025 equivalents) and the appropriatedihalo compound (2.5 equivalents) were added to a substituted phenylacetonitrile. The mixture was heated at 70° C. and then 50% sodiumhydroxide (10 equivalents) was slowly added to the mixture. The reactionwas stirred at 70° C. for 12-24 hours to ensure complete formation ofthe cycloalkyl moiety and then heated at 130° C. for 24-48 hours toensure complete conversion from the nitrile to the carboxylic acid. Thedark brown/black reaction mixture was diluted with water and extractedwith dichloromethane three times to remove side products. The basicaqueous solution was acidified with concentrated hydrochloric acid to pHless than one and the precipitate which began to form at pH 4 wasfiltered and washed with 1 M hydrochloric acid two times. The solidmaterial was dissolved in dichloromethane and extracted two times with 1M hydrochloric acid and one time with a saturated aqueous solution ofsodium chloride. The organic solution was dried over sodium sulfate andevaporated to dryness to give the cycloalkylcarboxylic acid. Yields andpurities were typically greater than 90%.

Example 1: 1-Benzo[1,3]dioxol-5-yl-cyclopropanecarboxylic Acid

A mixture of 2-(benzo[d][1,3]dioxol-5-yl)acetonitrile (5.10 g 31.7mmol), 1-bromo-2-chloro-ethane (9.00 mL 109 mmol), andbenzyltriethylammonium chloride (0.181 g, 0.795 mmol) was heated at 70°C. and then 50% (wt./wt.) aqueous sodium hydroxide (26 mL) was slowlyadded to the mixture. The reaction was stirred at 70° C. for 24 hoursand then heated at 130° C. for 48 hours. The dark brown reaction mixturewas diluted with water (400 mL) and extracted once with an equal volumeof ethyl acetate and once with an equal volume of dichloromethane. Thebasic aqueous solution was acidified with concentrated hydrochloric acidto pH less than one and the precipitate filtered and washed with 1 Mhydrochloric acid. The solid material was dissolved in dichloromethane(400 mL) and extracted twice with equal volumes of 1 M hydrochloric acidand once with a saturated aqueous solution of sodium chloride. Theorganic solution was dried over sodium sulfate and evaporated to drynessto give a white to slightly off-white solid (5.23 g, 80%) ESI-MS m zcalc. 206.1, found 207.1 (M+1)⁺. Retention time 2.37 minutes. ¹H NMR(400 MHz, DMSO-d₆) δ 1.07-1.11 (m, 2H), 1.38-1.42 (m, 2H), 5.98 (s, 2H),6.79 (m, 2H), 6.88 (m, 1H), 12.26 (s, 1H).

General Procedure II: Carboxylic Acid Building Block

Sodium hydroxide (50% aqueous solution, 7.4 equivalents) was slowlyadded to a mixture of the appropriate phenyl acetonitrile,benzyltriethylammonium chloride (1.1 equivalents), and the appropriatedihalo compound (2.3 equivalents) at 70° C. The mixture was stirredovernight at 70° C. and the reaction mixture was diluted with water (30mL) and extracted with ethyl acetate. The combined organic layers weredried over sodium sulfate and evaporated to dryness to give the crudecyclopropanecarbonitrile, which was used directly in the next step.

The crude cyclopropanecarbonitrile was refluxed in 10% aqueous sodiumhydroxide (7.4 equivalents) for 2.5 hours. The cooled reaction mixturewas washed with ether (100 mL) and the aqueous phase was acidified to pH2 with 2M hydrochloric acid. The precipitated solid was filtered to givethe cyclopropanecarboxylic acid as a white solid.

General Procedure III: Carboxylic Acid Building Block

Example 2: 1-(2,2-Difluoro-benzo[1,3]dioxol-5-yl)-cyclopropanecarboxylic

2,2-Difluoro-benzo[1,3]dioxole-5-carboxylic Acid Methyl Ester

A solution of 5-bromo-2,2-difluoro-benzo[1,3]dioxole (11.8 g, 50.0 mmol)and tetrakis(triphenylphosphine)palladium (0) [Pd(PPh₃)₄, 5.78 g, 5.00mmol] in methanol (20 mL) containing acetonitrile (30 mL) andtriethylamine (10 mL) was stirred under a carbon monoxide atmosphere (55PSI) at 75° C. (oil bath temperature) for 15 hours. The cooled reactionmixture was filtered and the filtrate was evaporated to dryness. Theresidue was purified by silica gel column chromatography to give crude2,2-difluoro-benzo [1,3] dioxole-5-carboxylic acid methyl ester (11.5g), which was used directly in the next step.

(2,2-Difluoro-benzo[1,3]dioxol-5-yl)-methanol

Crude 2,2-difluoro-benzo[1,3]dioxole-5-carboxylic acid methyl ester(11.5 g) dissolved in 20 mL of anhydrous tetrahydrofuran (THF) wasslowly added to a suspension of lithium aluminum hydride (4.10 g, 106mmol) in anhydrous THF (100 mL) at 0° C. The mixture was then warmed toroom temperature. After being stirred at room temperature for 1 hour,the reaction mixture was cooled to 0° C. and treated with water (4.1 g),followed by sodium hydroxide (10% aqueous solution, 4.1 mL). Theresulting slurry was filtered and washed with THF. The combined filtratewas evaporated to dryness and the residue was purified by silica gelcolumn chromatography to give(2,2-difluoro-benzo[1,3]dioxol-5-yl)-methanol (7.2 g, 38 mmol, 76% overtwo steps) as a colorless oil.

5-Chloromethyl-2,2-difluoro-benzo[1,3]dioxole

Thionyl chloride (45 g, 38 mmol) was slowly added to a solution of(2,2-difluoro-benzo[1,3]dioxol-5-yl)-methanol (7.2 g, 38 mmol) indichloromethane (200 mL) at 0° C. The resulting mixture was stirredovernight at room temperature and then evaporated to dryness. Theresidue was partitioned between an aqueous solution of saturated sodiumbicarbonate (100 mL) and dichloromethane (100 mL). The separated aqueouslayer was extracted with dichloromethane (150 mL) and the organic layerwas dried over sodium sulfate, filtrated, and evaporated to dryness togive crude 5-chloromethyl-2,2-difluoro-benzo[1,3]dioxole (4.4 g) whichwas used directly in the next step.

(2,2-Difluoro-benzo[1,3]dioxol-5-yl)-acetonitrile

A mixture of crude 5-chloromethyl-2,2-difluoro-benzo[1,3]dioxole (4.4 g)and sodium cyanide (1.36 g, 27.8 mmol) in dimethylsulfoxide (50 mL) wasstirred at room temperature overnight. The reaction mixture was pouredinto ice and extracted with ethyl acetate (300 mL). The organic layerwas dried over sodium sulfate and evaporated to dryness to give crude(2,2-difluoro-benzo[1,3]dioxol-5-yl)-acetonitrile (3.3 g) which was useddirectly in the next step.

1-(2,2-Difluoro-benzo[1,3]dioxol-5-yl)-cyclopropanecarbonitrile

Sodium hydroxide (50% aqueous solution, 10 mL) was slowly added to amixture of crude (2,2-difluoro-benzo[1,3]dioxol-5-yl)-acetonitrile,benzyltriethylammonium chloride (3.00 g, 15.3 mmol), and1-bromo-2-chloroethane (4.9 g, 38 mmol) at 70° C.

The mixture was stirred overnight at 70° C. before the reaction mixturewas diluted with water (30 mL) and extracted with ethyl acetate. Thecombined organic layers were dried over sodium sulfate and evaporated todryness to give crude1-(2,2-difluoro-benzo[1,3]dioxol-5-yl)-cyclopropanecarbonitrile, whichwas used directly in the next step.

1-(2,2-Difluoro-benzo[1,3]dioxol-5-yl)-cyclopropanecarboxylic Acid

1-(2,2-Difluoro-benzo[1,3]dioxol-5-yl)-cyclopropanecarbonitrile (crudefrom the last step) was refluxed in 10% aqueous sodium hydroxide (50 mL)for 2.5 hours. The cooled reaction mixture was washed with ether (100mL) and the aqueous phase was acidified to pH 2 with 2M hydrochloricacid. The precipitated solid was filtered to give1-(2,2-difluoro-benzo[1,3]dioxol-5-yl)-cyclopropanecarboxylic acid as awhite solid (0.15 g, 1.6% over four steps). ESI-MS m/z calc. 242.04,found 241.58 (M+1)⁺; ¹H NMR (CDCl₃) δ 7.14-7.04 (m, 2H), 6.98-6.96 (m,1H), 1.74-1.64 (m, 2H), 1.26-1.08 (m, 2H).

Example 3: 2-(2,2-Dimethylbenzo[d][1,3]dioxol-5-yl)acetonitrile

(3,4-Dihydroxy-phenyl)-acetonitrile

To a solution of benzo[1,3]dioxol-5-yl-acetonitrile (0.50 g, 3.1 mmol)in CH₂Cl₂ (15 mL) was added dropwise BBr₃ (0.78 g, 3.1 mmol) at −78° C.under N₂. The mixture was slowly warmed to room temperature and stirredovernight. H₂O (10 mL) was added to quench the reaction and the CH₂Cl₂layer was separated. The aqueous phase was extracted with CH₂Cl₂ (2×7mL). The combined organics were washed with brine, dried over Na₂SO₄ andpurified by column chromatography on silica gel (petroleum ether/ethylacetate 5:1) to give (3,4-dihydroxy-phenyl)-acetonitrile (0.25 g, 54%)as a white solid. ¹H NMR (DMSO-d₆, 400 MHz) δ 9.07 (s, 1H), 8.95 (s,1H), 6.68-6.70 (m, 2H), 6.55 (dd, J=8.0, 2.0 Hz, 1H), 3.32 (s, 2H).

2-(2,2-Dimethylbenzo[d][1,3]dioxol-5-yl)acetonitrile

To a solution of (3,4-dihydroxy-phenyl)-acetonitrile (0.20 g, 1.3 mmol)in toluene (4 mL) was added 2,2-dimethoxy-propane (0.28 g, 2.6 mmol) andTsOH (0.010 g, 0.065 mmol). The mixture was heated at reflux overnight.The reaction mixture was evaporated to remove the solvent and theresidue was dissolved in ethyl acetate. The organic layer was washedwith NaHCO₃ solution, H₂O, brine, and dried over Na₂SO₄. The solvent wasevaporated under reduced pressure to give a residue, which was purifiedby column chromatography on silica gel (petroleum ether/ethyl acetate10:1) to give 2-(2,2-dimethylbenzo[d][1,3]dioxol-5-yl)acetonitrile (40mg, 20%). ¹H NMR (CDCl₃, 400 MHz) δ 6.68-6.71 (m, 3H), 3.64 (s, 2H),1.67 (s, 6H).

Example 4: 1-(3,4-Dihydroxy-phenyl)-cyclopropanecarboxylic Acid

1-(3,4-Bis-benzyloxy-phenyl)-cyclopropanecarbonitrile

To a mixture of (n-C₄H9)₄NBr (0.50 g, 1.5 mmol), toluene (7 mL) and(3,4-bis-benzyloxy-phenyl)-acetonitrile (14 g, 42 mmol) in NaOH (50 g)and H₂O (50 mL) was added BrCH₂CH₂Cl (30 g, 0.21 mol). The reactionmixture was stirred at 50° C. for 5 h before being cooled to roomtemperature. Toluene (30 mL) was added and the organic layer wasseparated and washed with H₂O, brine, dried over anhydrous MgSO₄, andconcentrated. The residue was purified by column on silica gel(petroleum ether/ethyl acetate 10:1) to give1-(3,4-bis-benzyloxy-phenyl)-cyclopropanecarbonitrile (10 g, 66%). ¹HNMR (DMSO 300 MHz) δ 7.46-7.30 (m, 10H), 7.03 (d, J=8.4 Hz, 1H), 6.94(d, J=2.4 Hz, 1H), 6.89 (dd, J=2.4, 8.4 Hz, 1H), 5.12 (d, J=7.5 Hz, 4H),1.66-1.62 (m, 2H), 1.42-1.37 (m, 2H).

1-(3,4-Dihydroxy-phenyl)-cyclopropanecarbonitrile

To a solution of 1-(3,4-bis-benzyloxy-phenyl)-cyclopropanecarbonitrile(10 g, 28 mmol) in MeOH (50 mL) was added Pd/C (0.5 g) under nitrogenatmosphere. The mixture was stirred under hydrogen atmosphere (1 atm) atroom temperature for 4 h. The catalyst was filtered off through a celitepad and the filtrate was evaporated under vacuum to give1-(3,4-dihydroxy-phenyl)-cyclopropanecarbonitrile (4.5 g, 92%). ¹H NMR(DMSO 400 MHz) δ 9.06 (br s, 2H), 6.67-6.71 (m, 2H), 6.54 (dd, J=2.4,8.4 Hz, 1H), 1.60-1.57 (m, 2H), 1.30-1.27 (m, 2H)

1-(3,4-Dihydroxy-phenyl)-cyclopropanecarboxylic Acid

To a solution of NaOH (20 g, 0.50 mol) in H₂O (20 mL) was added1-(3,4-dihydroxy-phenyl)-cyclopropanecarbonitrile (4.4 g, 25 mmol). Themixture was heated at reflux for 3 h before being cooled to roomtemperature. The mixture was neutralized with HCl (0.5 N) to pH 3-4 andextracted with ethyl acetate (20 mL×3). The combined organic layers werewashed with water, brine, dried over anhydrous MgSO₄, and concentratedunder vacuum to obtain 1-(3,4-dihydroxy-phenyl)-cyclopropanecarboxylicacid (4.5 g crude). From 900 mg crude, 500 mg pure1-(3,4-dihydroxy-phenyl)-cyclopropanecarboxylic acid was obtained bypreparatory HPLC. ¹H NMR (DMSO, 300 MHz) δ 12.09 (br s, 1H), 8.75 (br s,2H), 6.50-6.67 (m, 3H), 1.35-1.31 (m, 2H), 1.01-0.97 (m, 2H).

Example 5:1-(2-Oxo-2,3-dihydrobenzo[d]oxazol-5-yl)cyclopropane-carboxylic Acid

1-(4-Methoxy-phenyl)-cyclopropanecarboxylic Acid Methyl Ester

To a solution of 1-(4-methoxy-phenyl)-cyclopropanecarboxylic acid (50 g,0.26 mol) in MeOH (500 mL) was added toluene-4-sulfonic acid monohydrate(2.5 g, 13 mmol) at room temperature. The reaction mixture was heated atreflux for 20 hours. MeOH was removed by evaporation under vacuum andEtOAc (200 mL) was added. The organic layer was washed with sat. aq.NaHCO₃ (100 mL) and brine, dried over anhydrous Na₂SO₄ and evaporatedunder vacuum to give 1-(4-methoxy-phenyl)-cyclopropanecarboxylic acidmethyl ester (53 g, 99%). ¹H NMR (CDCl₃, 400 MHz) δ 7.25-7.27 (m, 2H),6.85 (d, J=8.8 Hz, 2H), 3.80 (s, 3H), 3.62 (s, 3H), 1.58 (q, J=3.6 Hz,2H), 1.15 (q, J=3.6 Hz, 2H).

1-(4-Methoxy-3-nitro-phenyl)-cyclopropanecarboxylic Acid Methyl Ester

To a solution of 1-(4-methoxy-phenyl)-cyclopropanecarboxylic acid methylester (30.0 g, 146 mmol) in Ac₂O (300 mL) was added a solution of HNO₃(14.1 g, 146 mmol, 65%) in AcOH (75 mL) at 0° C. The reaction mixturewas stirred at 0˜ 5° C. for 3 h before aq. HCl (20%) was added dropwiseat 0° C. The resulting mixture was extracted with EtOAc (200 mL×3). Theorganic layer was washed with sat. aq. NaHCO₃ then brine, dried overanhydrous Na₂SO₄ and evaporated under vacuum to give1-(4-methoxy-3-nitro-phenyl)-cyclopropanecarboxylic acid methyl ester(36.0 g, 98%), which was directly used in the next step. ¹H NMR (CDCl₃300 MHz) δ 7.84 (d, J=2.1 Hz, 1H), 7.54 (dd, J=2.1, 8.7 Hz, 1H), 7.05(d, J=8.7 Hz, 1H), 3.97 (s, 3H), 3.65 (s, 3H), 1.68-1.64 (m, 2H),1.22-1.18 (m, 2H).

1-(4-Hydroxy-3-nitro-phenyl)-cyclopropanecarboxylic Acid Methyl Ester

To a solution of 1-(4-methoxy-3-nitro-phenyl)-cyclopropane-carboxylicacid methyl ester (10.0 g, 39.8 mmol) in CH₂Cl₂ (100 mL) was added BBr₃(12.0 g, 47.8 mmol) at −70° C. The mixture was stirred at −70° C. for 1hour, then allowed to warm to −30° C. and stirred at this temperaturefor 3 hours. Water (50 mL) was added dropwise at −20° C., and theresulting mixture was allowed to warm room temperature before it wasextracted with EtOAc (200 mL×3). The combined organic layers were driedover anhydrous Na₂SO₄ and evaporated under vacuum to give the crudeproduct, which was purified by column chromatography on silica gel(petroleum ether/ethyl acetate 15:1) to afford1-(4-hydroxy-3-nitro-phenyl)-cyclopropanecarboxylic acid methyl ester(8.3 g, 78%). ¹H NMR (CDCl₃, 400 MHz) δ 10.5 (s, 1H), 8.05 (d, J=2.4 Hz,1H), 7.59 (dd, J=2.0, 8.8 Hz, 1H), 7.11 (d, J=8.4 Hz, 1H), 3.64 (s, 3H),1.68-1.64 (m, 2H), 1.20-1.15 (m, 2H).

1-(3-Amino-4-hydroxy-phenyl)-cyclopropanecarboxylic Acid Methyl Ester

To a solution of 1-(4-hydroxy-3-nitro-phenyl)-cyclopropanecarboxylicacid methyl ester (8.3 g, 35 mmol) in MeOH (100 mL) was added RaneyNickel (0.8 g) under nitrogen atmosphere. The mixture was stirred underhydrogen atmosphere (1 atm) at 35° C. for 8 hours. The catalyst wasfiltered off through a Celite pad and the filtrate was evaporated undervacuum to give crude product, which was purified by columnchromatography on silica gel (petroleum ether/ethyl acetate 1:1) to give1-(3-amino-4-hydroxy-phenyl)-cyclopropanecarboxylic acid methyl ester(5.3 g, 74%). ¹H NMR (CDCl₃, 400 MHz) δ 6.77 (s, 1H), 6.64 (d, J=2.0 Hz,2H), 3.64 (s, 3H), 1.55-1.52 (m, 2H), 1.15-1.12 (m, 2H).

1-(2-Oxo-2,3-dihydro-benzooxazol-5-yl)-cyclopropanecarboxylic AcidMethyl Ester

To a solution of 1-(3-amino-4-hydroxy-phenyl)-cyclopropanecarboxylicacid methyl ester (2.0 g, 9.6 mmol) in THF (40 mL) was added triphosgene(4.2 g, 14 mmol) at room temperature. The mixture was stirred for 20minutes at this temperature before water (20 mL) was added dropwise at0° C. The resulting mixture was extracted with EtOAc (100 mL×3). Thecombined organic layers were dried over anhydrous Na₂SO₄ and evaporatedunder vacuum to give1-(2-oxo-2,3-dihydro-benzooxazol-5-yl)-cyclopropanecarboxylic acidmethyl ester (2.0 g, 91%), which was directly used in the next step. ¹HNMR (CDCl₃, 300 MHz) δ 8.66 (s, 1H), 7.13-7.12 (m, 2H), 7.07 (s, 1H),3.66 (s, 3H), 1.68-1.65 (m, 2H), 1.24-1.20 (m, 2H).

1-(2-Oxo-2,3-dihydrobenzo[d]oxazol-5-yl)cyclopropanecarboxylic Acid

To a solution of1-(2-oxo-2,3-dihydro-benzooxazol-5-yl)-cyclopropanecarboxylic acidmethyl ester (1.9 g, 8.1 mmol) in MeOH (20 mL) and water (2 mL) wasadded LiOH.H₂O (1.7 g, 41 mmol) in portions at room temperature. Thereaction mixture was stirred for 20 hours at 50° C. MeOH was removed byevaporation under vacuum before water (100 mL) and EtOAc (50 mL) wereadded. The aqueous layer was separated, acidified with HCl (3 mol/L) andextracted with EtOAc (100 mL×3). The combined organic layers were driedover anhydrous Na₂SO₄ and evaporated under vacuum to give1-(2-oxo-2,3-dihydrobenzo[d]oxazol-5-yl)cyclopropanecarboxylic acid (1.5g, 84%). ¹H NMR (DMSO, 400 MHz) δ 12.32 (brs, 1H), 11.59 (brs, 1H), 7.16(d, J=8.4 Hz, 1H), 7.00 (d, J=8.0 Hz, 1H), 1.44-1.41 (m, 2H), 1.13-1.10(m, 2H). MS (ESI) m/e (M+H⁺) 218.1.

Example 6: 1-(6-Fluoro-benzo[1,3]dioxol-5-yl)-cyclopropanecarboxylicAcid

2-Fluoro-4,5-dihydroxy-benzaldehyde

To a stirred suspension of 2-fluoro-4,5-dimethoxy-benzaldehyde (3.00 g,16.3 mmol) in dichloromethane (100 mL) was added BBr₃ (12.2 mL, 130mmol) dropwise at −78° C. under nitrogen atmosphere. After addition, themixture was warmed to −30° C. and stirred at this temperature for 5 h.The reaction mixture was poured into ice water and the precipitatedsolid was collected by filtration and washed with dichloromethane toafford 2-fluoro-4,5-dihydroxy-benzaldehyde (8.0 g), which was useddirectly in the next step.

6-Fluoro-benzo[1,3]dioxole-5-carbaldehyde

To a stirred solution of 2-fluoro-4,5-dihydroxy-benzaldehyde (8.0 g) andBrClCH₂ (24.8 g, 190 mmol) in dry DMF (50 mL) was added Cs₂CO₃ (62.0 g,190 mmol) in portions. The resulting mixture was stirred at 60° C.overnight and then poured into water. The mixture was extracted withEtOAc (200 mL×3). The combined organic layers were washed with brine(200 mL), dried over Na₂SO₄, and evaporated in vacuo to give crudeproduct, which was purified by column chromatography on silica gel(5-20% ethyl acetate/petroleum ether) to afford6-fluoro-benzo[1,3]dioxole-5-carbaldehyde (700 mg, two steps yield:24%). ¹H-NMR (400 MHz, CDCl₃) δ 10.19 (s, 1H), 7.23 (d, J=5.6, 1H), 6.63(d, J=9.6, 1H), 6.08 (s, 2H).

(6-Fluoro-benzo[1,3]dioxol-5-yl)-methanol

To a stirred solution of 6-fluoro-benzo[1,3]dioxole-5-carbaldehyde (700mg, 4.2 mmol) in MeOH (50 mL) was added NaBH₄ (320 mg, 8.4 mmol) inportions at 0° C. The mixture was stirred at this temperature for 30 minand was then concentrated in vacuo to give a residue. The residue wasdissolved in EtOAc and the organic layer was washed with water, driedover Na₂SO₄, and concentrated in vacuo to afford(6-fluoro-benzo[1,3]dioxol-5-yl)-methanol (650 mg, 92%), which wasdirectly used in the next step.

5-Chloromethyl-6-fluoro-benzo[1,3]dioxole

(6-Fluoro-benzo[1,3]dioxol-5-yl)-methanol (650 mg, 3.8 mmol) was addedto SOCl₂ (20 mL) in portions at 0° C. The mixture was warmed to roomtemperature for 1 h and then heated at reflux for 1 h. The excess SOCl₂was evaporated under reduced pressure to give the crude product, whichwas basified with sat. NaHCO₃ solution to pH˜7. The aqueous phase wasextracted with EtOAc (50 mL×3). The combined organic layers were driedover Na₂SO₄ and evaporated under reduced pressure to give5-chloromethyl-6-fluoro-benzo[1,3]dioxole (640 mg, 90%), which wasdirectly used in the next step.

(6-Fluoro-benzo[1,3]dioxol-5-yl)-acetonitrile

A mixture of 5-chloromethyl-6-fluoro-benzo[1,3]dioxole (640 mg, 3.4mmol) and NaCN (340 mg, 6.8 mmol) in DMSO (20 mL) was stirred at 30° C.for 1 h and then poured into water. The mixture was extracted with EtOAc(50 mL×3). The combined organic layers were washed with water (50 mL)and brine (50 mL), dried over Na₂SO₄, and evaporated under reducedpressure to give the crude product, which was purified by columnchromatography on silica gel (5-10% ethyl acetate/petroleum ether) toafford (6-fluoro-benzo[1,3]dioxol-5-yl)-acetonitrile (530 mg, 70%).¹H-NMR (300 MHz, CDCl₃) δ 6.82 (d, J=4.8, 1H), 6.62 (d, J=5.4, 1H), 5.99(s, 2H), 3.65 (s, 2H).

1-(6-Fluoro-benzo[1,3]dioxol-5-yl)-cyclopropanecarbonitrile

A flask was charged with water (10 mL), followed by a rapid addition ofNaOH (10 g, 0.25 mol) in three portions over a 5 min period. The mixturewas allowed to cool to room temperature. Subsequently, the flask wascharged with toluene (6 mL), tetrabutyl-ammonium bromide (50 mg, 0.12mmol), (6-fluoro-benzo[1,3]dioxol-5-yl)-acetonitrile (600 mg, 3.4 mmol)and 1-bromo-2-chloroethane (1.7 g, 12 mmol). The mixture stirredvigorously at 50° C. overnight. The cooled flask was charged withadditional toluene (20 mL). The organic layer was separated and washedwith water (30 mL) and brine (30 mL). The organic layer was removed invacuo to give the crude product, which was purified by columnchromatography on silica gel (5-10% ethyl acetate/petroleum ether) togive 1-(6-fluoro-benzo[1,3]dioxol-5-yl)-cyclopropanecarbonitrile (400mg, 60%). ¹H NMR (300 MHz, CDCl₃) δ 6.73 (d, J=3.0 Hz, 1H), 6.61 (d,J=9.3 Hz, 1H), 5.98 (s, 2H), 1.67-1.62 (m, 2H), 1.31-1.27 (m, 2H).

1-(6-Fluoro-benzo[1,3]dioxol-5-yl)-cyclopropanecarboxylic acid

A mixture of 1-(6-fluoro-benzo[1,3]dioxol-5-yl)-cyclopropanecarbonitrile(400 mg, 0.196 mmol) and 10% NaOH (10 mL) was stirred at 100° C.overnight. After the reaction was cooled, 5% HCl was added until thepH<5 and then EtOAc (30 mL) was added to the reaction mixture. Thelayers were separated and combined organic layers were evaporated invacuo to afford1-(6-fluoro-benzo[1,3]dioxol-5-yl)-cyclopropanecarboxylic acid (330 mg,76%). ¹H NMR (400 MHz, DMSO) δ 12.2 (s, 1H), 6.87-6.85 (m, 2H), 6.00 (s,1H), 1.42-1.40 (m, 2H), 1.14-1.07 (m, 2H).

Example 7: 1-(Benzofuran-5-yl)cyclopropanecarboxylic Acid

1-[4-(2,2-Diethoxy-ethoxy)-phenyl]-cyclopropanecarboxylic Acid

To a stirred solution of 1-(4-hydroxy-phenyl)-cyclopropanecarboxylicacid methyl ester (15.0 g, 84.3 mmol) in DMF (50 mL) was added sodiumhydride (6.7 g, 170 mmol, 60% in mineral oil) at 0° C. After hydrogenevolution ceased, 2-bromo-1,1-diethoxy-ethane (16.5 g, 84.3 mmol) wasadded dropwise to the reaction mixture. The reaction was stirred at 160°C. for 15 hours. The reaction mixture was poured onto ice (100 g) andwas extracted with CH₂Cl₂. The combined organics were dried over Na₂SO₄.The solvent was evaporated under vacuum to give1-[4-(2,2-diethoxy-ethoxy)-phenyl]-cyclopropanecarboxylic acid (10 g),which was used directly in the next step without purification.

1-Benzofuran-5-yl-cyclopropanecarboxylic Acid

To a suspension of1-[4-(2,2-diethoxy-ethoxy)-phenyl]-cyclopropanecarboxylic acid (20 g,˜65 mmol) in xylene (100 mL) was added PPA (22.2 g, 64.9 mmol) at roomtemperature. The mixture was heated at reflux (140° C.) for 1 hourbefore it was cooled to room temperature and decanted from the PPA. Thesolvent was evaporated under vacuum to obtain the crude product, whichwas purified by preparative HPLC to provide1-(benzofuran-5-yl)cyclopropanecarboxylic acid (1.5 g, 5%). ¹H NMR (400MHz, DMSO-d₆) δ 12.25 (br s, 1H), 7.95 (d, J=2.8 Hz, 1H), 7.56 (d, J=2.0Hz, 1H), 7.47 (d, J=11.6 Hz, 1H), 7.25 (dd, J=2.4, 11.2 Hz, 1H), 6.89(d, J=1.6 Hz, 1H), 1.47-1.44 (m, 2H), 1.17-1.14 (m, 2H).

Example 8: 1-(2,3-Dihydrobenzofuran-6-yl)cyclopropanecarboxylic Acid

To a solution of 1-(benzofuran-6-yl)cyclopropanecarboxylic acid (370 mg,1.8 mmol) in MeOH (50 mL) was added PtO₂ (75 mg, 20%) at roomtemperature. The reaction mixture was stirred under hydrogen atmosphere(1 atm) at 20° C. for 3 d. The reaction mixture was filtered and thesolvent was evaporated in vacuo to afford the crude product, which waspurified by prepared HPLC to give1-(2,3-dihydrobenzofuran-6-yl)cyclopropanecarboxylic acid (155 mg, 42%).¹H NMR (300 MHz, MeOD) δ 7.13 (d, J=7.5 Hz, 1H), 6.83 (d, J=7.8 Hz, 1H),6.74 (s, 1H), 4.55 (t, J=8.7 Hz, 2H), 3.18 (t, J=8.7 Hz, 2H), 1.56-1.53(m, 2H), 1.19-1.15 (m, 2H).

Example 9:1-(3,3-Dimethyl-2,3-dihydrobenzofuran-5-yl)cyclopropanecarboxylic Acid

1-(4-Hydroxy-phenyl)-cyclopropanecarboxylic Acid Methyl Ester

To a solution of methyl 1-(4-methoxyphenyl)cyclopropanecarboxylate (10.0g, 48.5 mmol) in dichloromethane (80 mL) was added EtSH (16 mL) underice-water bath. The mixture was stirred at 0° C. for 20 min before AlCl₃(19.5 g, 0.15 mmol) was added slowly at 0° C. The mixture was stirred at0° C. for 30 min. The reaction mixture was poured into ice-water, theorganic layer was separated, and the aqueous phase was extracted withdichloromethane (50 mL×3). The combined organic layers were washed withH₂O, brine, dried over Na₂SO₄ and evaporated under vacuum to give1-(4-hydroxy-phenyl)-cyclopropanecarboxylic acid methyl ester (8.9 g,95%). ¹H NMR (400 MHz, CDCl₃) δ 7.20-7.17 (m, 2H), 6.75-6.72 (m, 2H),5.56 (s, 1H), 3.63 (s, 3H), 1.60-1.57 (m, 2H), 1.17-1.15 (m, 2H).

1-(4-Hydroxy-3,5-diiodo-phenyl)-cyclopropanecarboxylic Acid Methyl Ester

To a solution of 1-(4-hydroxy-phenyl)-cyclopropanecarboxylic acid methylester (8.9 g, 46 mmol) in CH₃CN (80 mL) was added NIS (15.6 g, 69 mmol).The mixture was stirred at room temperature for 1 hour. The reactionmixture was concentrated and the residue was purified by columnchromatography on silica gel (petroleum ether/ethyl acetate 10:1) togive 1-(4-hydroxy-3,5-diiodo-phenyl)-cyclopropanecarboxylic acid methylester (3.5 g, 18%). ¹H NMR (400 MHz, CDCl₃) δ 7.65 (s, 2H), 5.71 (s,1H), 3.63 (s, 3H), 1.59-1.56 (m, 2H), 1.15-1.12 (m, 2H).

1-[3,5-Diiodo-4-(2-methyl-allyloxy)-phenyl]-cyclopropanecarboxylic AcidMethyl Ester

A mixture of 1-(4-hydroxy-3,5-diiodo-phenyl)-cyclopropanecarboxylic acidmethyl ester (3.2 g, 7.2 mmol), 3-chloro-2-methyl-propene (1.0 g, 11mmol), K₂CO₃ (1.2 g, 8.6 mmol), NaI (0.1 g, 0.7 mmol) in acetone (20 mL)was stirred at 20° C. overnight. The solid was filtered off and thefiltrate was concentrated under vacuum to give1-[3,5-diiodo-4-(2-methyl-allyloxy)-phenyl]-cyclopropane-carboxylic acidmethyl ester (3.5 g, 97%). ¹H NMR (300 MHz, CDCl₃) δ 7.75 (s, 2H), 5.26(s, 1H), 5.06 (s, 1H), 4.38 (s, 2H), 3.65 (s, 3H), 1.98 (s, 3H),1.62-1.58 (m, 2H), 1.18-1.15 (m, 2H).

1-(3,3-Dimethyl-2,3-dihydro-benzofuran-5-yl)-cyclopropanecarboxylic AcidMethyl Ester

To a solution of1-[3,5-diiodo-4-(2-methyl-allyloxy)-phenyl]-cyclopropane-carboxylic acidmethyl ester (3.5 g, 7.0 mmol) in toluene (15 mL) was added Bu₃SnH (2.4g, 8.4 mmol) and AIBN (0.1 g, 0.7 mmol). The mixture was heated atreflux overnight. The reaction mixture was concentrated under vacuum andthe residue was purified by column chromatography on silica gel(petroleum ether/ethyl acetate 20:1) to give1-(3,3-dimethyl-2,3-dihydro-benzofuran-5-yl)-cyclopropanecarboxylic acidmethyl ester (1.05 g, 62%). ¹H NMR (400 MHz, CDCl₃) δ 7.10-7.07 (m, 2H),6.71 (d, J=8 Hz, 1H), 4.23 (s, 2H), 3.62 (s, 3H), 1.58-1.54 (m, 2H),1.34 (s, 6H), 1.17-1.12 (m, 2H).

1-(3,3-Dimethyl-2,3-dihydrobenzofuran-5-yl)cyclopropanecarboxylic Acid

To a solution of1-(3,3-dimethyl-2,3-dihydro-benzofuran-5-yl)-cyclopropanecarboxylic acidmethyl ester (1.0 g, 4.0 mmol) in MeOH (10 mL) was added LiOH (0.40 g,9.5 mmol). The mixture was stirred at 40° C. overnight. HCl (10%) wasadded slowly to adjust the pH to 5. The resulting mixture was extractedwith ethyl acetate (10 mL×3). The extracts were washed with brine anddried over Na₂SO₄. The solvent was removed under vacuum and the crudeproduct was purified by preparative HPLC to give1-(3,3-dimethyl-2,3-dihydrobenzofuran-5-yl)cyclopropanecarboxylic acid(0.37 g, 41%). ¹H NMR (400 MHz, CDCl₃) δ 7.11-7.07 (m, 2H), 6.71 (d, J=8Hz, 1H), 4.23 (s, 2H), 1.66-1.63 (m, 2H), 1.32 (s, 6H), 1.26-1.23 (m,2H).

Example 10: 2-(7-Methoxybenzo[d][1,3]dioxol-5-yl)acetonitrile

3,4-Dihydroxy-5-methoxybenzoate

To a solution of 3,4,5-trihydroxy-benzoic acid methyl ester (50 g, 0.27mol) and Na₂B₄O₇ (50 g) in water (1000 mL) was added Me₂SO₄ (120 mL) andaqueous NaOH solution (25%, 200 mL) successively at room temperature.The mixture was stirred at room temperature for 6 h before it was cooledto 0° C. The mixture was acidified to pH˜2 by adding conc. H₂SO₄ andthen filtered. The filtrate was extracted with EtOAc (500 mL×3). Thecombined organic layers were dried over anhydrous Na₂SO₄ and evaporatedunder reduced pressure to give methyl 3,4-dihydroxy-5-methoxybenzoate(15.3 g 47%), which was used in the next step without furtherpurification.

Methyl 7-methoxybenzo[d][1,3]dioxole-5-carboxylate

To a solution of methyl 3,4-dihydroxy-5-methoxybenzoate (15.3 g, 0.0780mol) in acetone (500 mL) was added CH₂BrCl (34.4 g, 0.270 mol) and K₂CO₃(75.0 g, 0.540 mol) at 80° C. The resulting mixture was heated at refluxfor 4 h. The mixture was cooled to room temperature and solid K₂CO₃ wasfiltered off. The filtrate was concentrated under reduced pressure, andthe residue was dissolved in EtOAc (100 mL). The organic layer waswashed with water, dried over anhydrous Na₂SO₄, and evaporated underreduced pressure to give the crude product, which was purified by columnchromatography on silica gel (petroleum ether/ethyl acetate=10:1) toafford methyl 7-methoxybenzo[d][1,3]dioxole-5-carboxylate (12.6 g, 80%).¹H NMR (400 MHz, CDCl₃) δ 7.32 (s, 1H), 7.21 (s, 1H), 6.05 (s, 2H), 3.93(s, 3H), 3.88 (s, 3H).

(7-Methoxybenzo[d][1,3]dioxol-5-yl)methanol

To a solution of methyl 7-methoxybenzo[d][1,3]dioxole-5-carboxylate (14g, 0.040 mol) in THF (100 mL) was added LiAlH₄ (3.1 g, 0.080 mol) inportions at room temperature. The mixture was stirred for 3 h at roomtemperature. The reaction mixture was cooled to 0° C. and treated withwater (3.1 g) and NaOH (10%, 3.1 mL) successively. The slurry wasfiltered off and washed with THF. The combined filtrates were evaporatedunder reduced pressure to give(7-methoxy-benzo[d][1,3]dioxol-5-yl)methanol (7.2 g, 52%). ¹H NMR (400MHz, CDCl₃) δ 6.55 (s, 1H), 6.54 (s, 1H), 5.96 (s, 2H), 4.57 (s, 2H),3.90 (s, 3H).

6-(Chloromethyl)-4-methoxybenzo[d][1,3]dioxole

To a solution of SOCl₂ (150 mL) was added(7-methoxybenzo[d][1,3]dioxol-5-yl)methanol (9.0 g, 54 mmol) in portionsat 0° C. The mixture was stirred for 0.5 h. The excess SOCl₂ wasevaporated under reduced pressure to give the crude product, which wasbasified with sat. aq. NaHCO₃ to pH˜7. The aqueous phase was extractedwith EtOAc (100 mL×3). The combined organic layers were dried overanhydrous Na₂SO₄ and evaporated to give6-(chloromethyl)-4-methoxybenzo[d][1,3]dioxole (10 g 94%), which wasused in the next step without further purification. ¹H NMR (400 MHz,CDCl₃) δ 6.58 (s, 1H), 6.57 (s, 1H), 5.98 (s, 2H), 4.51 (s, 2H), 3.90(s, 3H).

2-(7-Methoxybenzo[d][1,3]dioxol-5-yl)acetonitrile

To a solution of 6-(chloromethyl)-4-methoxybenzo[d][1,3]dioxole (10 g,40 mmol) in DMSO (100 mL) was added NaCN (2.4 g, 50 mmol) at roomtemperature. The mixture was stirred for 3 h and poured into water (500mL). The aqueous phase was extracted with EtOAc (100 mL×3). The combinedorganic layers were dried over anhydrous Na₂SO₄ and evaporated to givethe crude product, which was washed with ether to afford2-(7-methoxybenzo[d][1,3]dioxol-5-yl)acetonitrile (4.6 g, 45%). ¹H NMR(400 MHz, CDCl₃) δ 6.49 (s, 2H), 5.98 (s, 2H), 3.91 (s, 3H), 3.65 (s,2H). ¹³C NMR (400 MHz, CDCl₃) δ 148.9, 143.4, 134.6, 123.4, 117.3,107.2, 101.8, 101.3, 56.3, 23.1.

Example 11: 2-(3-(Benzyloxy)-4-methoxyphenyl)acetonitrile

To a suspension of t-BuOK (20.2 g, 0.165 mol) in THF (250 mL) was addeda solution of TosMIC (16.1 g, 82.6 mmol) in THF (100 mL) at −78° C. Themixture was stirred for 15 minutes, treated with a solution of3-benzyloxy-4-methoxy-benzaldehyde (10.0 g, 51.9 mmol) in THF (50 mL)dropwise, and continued to stir for 1.5 hours at −78° C. To the cooledreaction mixture was added methanol (50 mL). The mixture was heated atreflux for 30 minutes. Solvent was removed to give a crude product,which was dissolved in water (300 mL). The aqueous phase was extractedwith EtOAc (100 mL×3). The combined organic layers were dried andevaporated under reduced pressure to give crude product, which waspurified by column chromatography (petroleum ether/ethyl acetate 10:1)to afford 2-(3-(benzyloxy)-4-methoxyphenyl)-acetonitrile (5.0 g, 48%).¹H NMR (300 MHz, CDCl₃) δ 7.48-7.33 (m, 5H), 6.89-6.86 (m, 3H), 5.17 (s,2H), 3.90 (s, 3H), 3.66 (s, 2H). ¹³C NMR (75 MHz, CDCl₃) δ 149.6, 148.6,136.8, 128.8, 128.8, 128.2, 127.5, 127.5, 122.1, 120.9, 118.2, 113.8,112.2, 71.2, 56.2, 23.3.

Example 12: 2-(3-(Benzyloxy)-4-chlorophenyl)acetonitrile

(4-Chloro-3-hydroxy-phenyl)acetonitrile

BBr₃ (17 g, 66 mmol) was slowly added to a solution of2-(4-chloro-3-methoxyphenyl)acetonitrile (12 g, 66 mmol) indichloromethane (120 mL) at −78° C. under N₂. The reaction temperaturewas slowly increased to room temperature. The reaction mixture wasstirred overnight and then poured into ice and water. The organic layerwas separated, and the aqueous layer was extracted with dichloromethane(40 mL×3). The combined organic layers were washed with water, brine,dried over Na₂SO₄, and concentrated under vacuum to give(4-chloro-3-hydroxy-phenyl)-acetonitrile (9.3 g, 85%). ¹H NMR (300 MHz,CDCl₃) δ 7.34 (d, J=8.4 Hz, 1H), 7.02 (d, J=2.1 Hz, 1H), 6.87 (dd,J=2.1, 8.4 Hz, 1H), 5.15 (brs, 1H), 3.72 (s, 2H).

2-(3-(Benzyloxy)-4-chlorophenyl)acetonitrile

To a solution of (4-chloro-3-hydroxy-phenyl)acetonitrile (6.2 g, 37mmol) in CH₃CN (80 mL) was added K₂CO₃ (10 g, 74 mmol) and BnBr (7.6 g,44 mmol). The mixture was stirred at room temperature overnight. Thesolids were filtered off and the filtrate was evaporated under vacuum.The residue was purified by column chromatography on silica gel(petroleum ether/ethyl acetate 50:1) to give2-(3-(benzyloxy)-4-chlorophenyl)-acetonitrile (5.6 g, 60%). ¹H NMR (400MHz, CDCl₃) δ 7.48-7.32 (m, 6H), 6.94 (d, J=2 Hz, 2H), 6.86 (dd, J=2.0,8.4 Hz, 1H), 5.18 (s, 2H), 3.71 (s, 2H).

Example 13: 2-(3-(Benzyloxy)-4-methoxyphenyl)acetonitrile

To a suspension of t-BuOK (20.2 g, 0.165 mol) in THF (250 mL) was addeda solution of TosMIC (16.1 g, 82.6 mmol) in THF (100 mL) at −78° C. Themixture was stirred for 15 minutes, treated with a solution of3-benzyloxy-4-methoxy-benzaldehyde (10.0 g, 51.9 mmol) in THF (50 mL)dropwise, and continued to stir for 1.5 hours at −78° C. To the cooledreaction mixture was added methanol (50 mL). The mixture was heated atreflux for 30 minutes. Solvent of the reaction mixture was removed togive a crude product, which was dissolved in water (300 mL). The aqueousphase was extracted with EtOAc (100 mL×3). The combined organic layerswere dried and evaporated under reduced pressure to give crude product,which was purified by column chromatography (petroleum ether/ethylacetate 10:1) to afford 2-(3-(benzyloxy)-4-methoxyphenyl)acetonitril(5.0 g, 48%). ¹H NMR (300 MHz, CDCl₃) δ 7.48-7.33 (m, 5H), 6.89-6.86 (m,3H), 5.17 (s, 2H), 3.90 (s, 3H), 3.66 (s, 2H). ¹³C NMR (75 MHz, CDCl₃) δ149.6, 148.6, 136.8, 128.8, 128.8, 128.2, 127.5, 127.5, 122.1, 120.9,118.2, 113.8, 112.2, 71.2, 56.2, 23.3.

Example 14: 2-(3-Chloro-4-methoxyphenyl)acetonitrile

To a suspension of t-BuOK (4.8 g, 40 mmol) in THF (30 mL) was added asolution of TosMIC (3.9 g, 20 mmol) in THF (10 mL) at −78° C. Themixture was stirred for 10 minutes, treated with a solution of3-chloro-4-methoxy-benzaldehyde (1.7 g, 10 mmol) in THF (10 mL)dropwise, and continued to stir for 1.5 hours at −78° C. To the cooledreaction mixture was added methanol (10 mL). The mixture was heated atreflux for 30 minutes. Solvent of the reaction mixture was removed togive a crude product, which was dissolved in water (20 mL). The aqueousphase was extracted with EtOAc (20 mL×3). The combined organic layerswere dried and evaporated under reduced pressure to give crude product,which was purified by column chromatography (petroleum ether/ethylacetate 10:1) to afford 2-(3-chloro-4-methoxyphenyl)acetonitrile (1.5 g,83%). ¹H NMR (400 MHz, CDCl₃) δ 7.33 (d, J=2.4 Hz, 1H), 7.20 (dd, J=2.4,8.4 Hz, 1H), 6.92 (d, J=8.4 Hz, 1H), 3.91 (s, 3H), 3.68 (s, 2H). ¹³C NMR(100 MHz, CDCl₃) δ 154.8, 129.8, 127.3, 123.0, 122.7, 117.60, 112.4,56.2, 22.4.

Example 15: 2-(3-Fluoro-4-methoxyphenyl)acetonitrile

To a suspension of t-BuOK (25.3 g, 0.207 mol) in THF (150 mL) was addeda solution of TosMIC (20.3 g, 0.104 mol) in THF (50 mL) at −78° C. Themixture was stirred for 15 minutes, treated with a solution of3-fluoro-4-methoxy-benzaldehyde (8.00 g, 51.9 mmol) in THF (50 mL)dropwise, and continued to stir for 1.5 hours at −78° C. To the cooledreaction mixture was added methanol (50 mL). The mixture was heated atreflux for 30 minutes. Solvent of the reaction mixture was removed togive a crude product, which was dissolved in water (200 mL). The aqueousphase was extracted with EtOAc (100 mL×3). The combined organic layerswere dried and evaporated under reduced pressure to give crude product,which was purified by column chromatography (petroleum ether/ethylacetate 10:1) to afford 2-(3-fluoro-4-methoxyphenyl)acetonitrile (5.0 g,58%). ¹H NMR (400 MHz, CDCl₃) δ 7.02-7.05 (m, 2H), 6.94 (t, J=8.4 Hz,1H), 3.88 (s, 3H), 3.67 (s, 2H). ¹³C NMR (100 MHz, CDCl₃) δ 152.3,147.5, 123.7, 122.5, 117.7, 115.8, 113.8, 56.3, 22.6.

Example 16: 2-(4-Chloro-3-methoxyphenyl)acetonitrile

Chloro-2-methoxy-4-methyl-benzene

To a solution of 2-chloro-5-methyl-phenol (93 g, 0.65 mol) in CH₃CN (700mL) was added CH₃I (110 g, 0.78 mol) and K₂CO₃ (180 g, 1.3 mol). Themixture was stirred at 25° C. overnight. The solid was filtered off andthe filtrate was evaporated under vacuum to give1-chloro-2-methoxy-4-methyl-benzene (90 g, 89%). ¹H NMR (300 MHz, CDCl₃)δ 7.22 (d, J=7.8 Hz, 1H), 6.74-6.69 (m, 2H), 3.88 (s, 3H), 2.33 (s, 3H).

4-Bromomethyl-1-chloro-2-methoxy-benzene

To a solution of 1-chloro-2-methoxy-4-methyl-benzene (50 g, 0.32 mol) inCCl₄ (350 mL) was added NBS (57 g, 0.32 mol) and AIBN (10 g, 60 mmol).The mixture was heated at reflux for 3 hours. The solvent was evaporatedunder vacuum and the residue was purified by column chromatography onsilica gel (petroleum ether/ethyl acetate=20:1) to give4-bromomethyl-1-chloro-2-methoxy-benzene (69 g, 92%). ¹H NMR (400 MHz,CDCl₃) δ 7.33-7.31 (m, 1H), 6.95-6.91 (m, 2H), 4.46 (s, 2H), 3.92 (s,3H).

2-(4-Chloro-3-methoxyphenyl)acetonitrile

To a solution of 4-bromomethyl-1-chloro-2-methoxy-benzene (68.5 g, 0.290mol) in C₂H₅OH (90%, 500 mL) was added NaCN (28.5 g, 0.580 mol). Themixture was stirred at 60° C. overnight. Ethanol was evaporated and theresidue was dissolved in H₂O. The mixture was extracted with ethylacetate (300 mL×3). The combined organic layers were washed with brine,dried over Na₂SO₄ and purified by column chromatography on silica gel(petroleum ether/ethyl acetate 30:1) to give2-(4-chloro-3-methoxyphenyl)acetonitrile (25 g, 48%). ¹H NMR (400 MHz,CDCl₃) δ 7.36 (d, J=8 Hz, 1H), 6.88-6.84 (m, 2H), 3.92 (s, 3H), 3.74 (s,2H). ¹³C NMR (100 MHz, CDCl₃) δ 155.4, 130.8, 129.7, 122.4, 120.7,117.5, 111.5, 56.2, 23.5.

Example 17: 1-(3-(Hydroxymethyl)-4-methoxyphenyl)cyclopropanecarboxylicAcid

1-(4-Methoxy-phenyl)-cyclopropanecarboxylic Acid Methyl Ester

To a solution of 1-(4-methoxy-phenyl)-cyclopropanecarboxylic acid (50 g,0.26 mol) in MeOH (500 mL) was added toluene-4-sulfonic acid monohydrate(2.5 g, 13 mmol) at room temperature. The reaction mixture was heated atreflux for 20 hours. MeOH was removed by evaporation under vacuum andEtOAc (200 mL) was added. The organic layer was washed with sat. aq.NaHCO₃ (100 mL) and brine, dried over anhydrous Na₂SO₄ and evaporatedunder vacuum to give 1-(4-methoxy-phenyl)-cyclopropanecarboxylic acidmethyl ester (53 g, 99%). ¹H NMR (CDCl₃, 400 MHz) δ 7.25-7.27 (m, 2H),6.85 (d, J=8.8 Hz, 2H), 3.80 (s, 3H), 3.62 (s, 3H), 1.58 (m, 2H), 1.15(m, 2H).

1-(3-Chloromethyl-4-methoxy-phenyl)-cyclopropanecarboxylic Acid MethylEster

To a solution of 1-(4-methoxy-phenyl)-cyclopropanecarboxylic acid methylester (30.0 g, 146 mmol) and MOMCl (29.1 g, 364 mmol) in CS₂ (300 mL)was added TiCl₄ (8.30 g, 43.5 mmol) at 5° C. The reaction mixture washeated at 30° C. for 1 d and poured into ice-water. The mixture wasextracted with CH₂Cl₂ (150 mL×3). The combined organic extracts wereevaporated under vacuum to give1-(3-chloromethyl-4-methoxy-phenyl)-cyclopropanecarboxylic acid methylester (38.0 g), which was used in the next step without furtherpurification.

1-(3-Hydroxymethyl-4-methoxy-phenyl)-cyclopropanecarboxylic Acid MethylEster

To a suspension of1-(3-chloromethyl-4-methoxy-phenyl)-cyclopropanecarboxylic acid methylester (20 g) in water (350 mL) was added Bu₄NBr (4.0 g) and Na₂CO₃ (90g, 0.85 mol) at room temperature. The reaction mixture was heated at 65°C. overnight. The resulting solution was acidified with aq. HCl (2mol/L) and extracted with EtOAc (200 mL×3). The organic layer was washedwith brine, dried over anhydrous Na₂SO₄ and evaporated under vacuum togive crude product, which was purified by column (petroleum ether/ethylacetate 15:1) to give1-(3-hydroxymethyl-4-methoxy-phenyl)-cyclopropanecarboxylic acid methylester (8.0 g, 39%). ¹H NMR (CDCl₃, 400 MHz) δ 7.23-7.26 (m, 2H), 6.83(d, J=8.0 Hz, 1H), 4.67 (s, 2H), 3.86 (s, 3H), 3.62 (s, 3H), 1.58 (q,J=3.6 Hz, 2H), 1.14-1.17 (m, 2H).

1-[3-(tert-Butyl-dimethyl-silanyloxymethyl)-4-methoxy-phenyl]cyclopropaneCarboxylic Acid Methyl Ester

To a solution of1-(3-hydroxymethyl-4-methoxy-phenyl)-cyclopropanecarboxylic acid methylester (8.0 g, 34 mmol) in CH₂Cl₂ (100 mL) were added imidazole (5.8 g,85 mmol) and TBSCl (7.6 g, 51 mmol) at room temperature. The mixture wasstirred overnight at room temperature. The mixture was washed withbrine, dried over anhydrous Na₂SO₄ and evaporated under vacuum to givecrude product, which was purified by column (petroleum ether/ethylacetate 30:1) to give1-[3-(tert-butyl-dimethyl-silanyloxymethyl)-4-methoxy-phenyl]-cyclopropanecarboxylicacid methyl ester (6.7 g, 56%). ¹H NMR (CDCl₃, 400 MHz) δ 7.44-7.45 (m,1H), 7.19 (dd, J=2.0, 8.4 Hz, 1H), 6.76 (d, J=8.4 Hz, 1H), 4.75 (s, 2H),3.81 (s, 3H), 3.62 (s, 3H), 1.57-1.60 (m, 2H), 1.15-1.18 (m, 2H), 0.96(s, 9H), 0.11 (s, 6H).

1-(3-Hydroxymethyl-4-methoxy-phenyl)-cyclopropanecarboxylic Acid

To a solution of1-[3-(tert-butyl-dimethyl-silanyloxymethyl)-4-methoxy-phenyl]-cyclopropanecarboxylic acid methyl ester (6.2 g, 18 mmol) in MeOH (75 mL) was addeda solution of LiOH.H₂O (1.5 g, 36 mmol) in water (10 mL) at 0° C. Thereaction mixture was stirred overnight at 40° C. MeOH was removed byevaporation under vacuum. AcOH (1 mol/L, 40 mL) and EtOAc (200 mL) wereadded. The organic layer was separated, washed with brine, dried overanhydrous Na₂SO₄ and evaporated under vacuum to provide1-(3-hydroxymethyl-4-methoxy-phenyl)-cyclopropanecarboxylic acid (5.3g).

Example 18: 2-(7-Chlorobenzo[d][1,3]dioxol-5-yl)acetonitrile

3-Chloro-4,5-dihydroxybenzaldehyde

To a suspension of 3-chloro-4-hydroxy-5-methoxy-benzaldehyde (10 g, 54mmol) in dichloromethane (300 mL) was added BBr₃ (26.7 g, 107 mmol)dropwise at −40° C. under N₂. After addition, the mixture was stirred atthis temperature for 5 h and then was poured into ice water. Theprecipitated solid was filtered and washed with petroleum ether. Thefiltrate was evaporated under reduced pressure to afford3-chloro-4,5-dihydroxybenzaldehyde (9.8 g, 89%), which was directly usedin the next step.

7-Chlorobenzo[d][1,3]dioxole-5-carbaldehyde

To a solution of 3-chloro-4,5-dihydroxybenzaldehyde (8.0 g, 46 mmol) andBrClCH₂ (23.9 g, 185 mmol) in dry DMF (100 mL) was added Cs₂CO₃ (25 g,190 mmol). The mixture was stirred at 60° C. overnight and was thenpoured into water. The resulting mixture was extracted with EtOAc (50mL×3). The combined extracts were washed with brine (100 mL), dried overNa₂SO₄ and concentrated under reduced pressure to afford7-chlorobenzo[d][1,3]dioxole-5-carbaldehyde (6.0 g, 70%). ¹H NMR (400MHz, CDCl₃) δ 9.74 (s, 1H), 7.42 (d, J=0.4 Hz, 1H), 7.26 (d, J=3.6 Hz,1H), 6.15 (s, 2H).

(7-Chlorobenzo[d][1,3]dioxol-5-yl)methanol

To a solution of 7-chlorobenzo[d][1,3]dioxole-5-carbaldehyde (6.0 g, 33mmol) in THF (50 mL) was added NaBH₄ (2.5 g, 64 mmol)) in portions at 0°C. The mixture was stirred at this temperature for 30 min and thenpoured into aqueous NH₄Cl solution. The organic layer was separated, andthe aqueous phase was extracted with EtOAc (50 mL×3). The combinedextracts were dried over Na₂SO₄ and evaporated under reduced pressure toafford (7-chlorobenzo[d][1,3]dioxol-5-yl)methanol, which was directlyused in the next step.

4-Chloro-6-(chloromethyl)benzo[d][1,3]dioxole

A mixture of (7-chlorobenzo[d][1,3]-dioxol-5-yl)methanol (5.5 g, 30mmol) and SOCl₂ (5.0 mL, 67 mmol) in dichloromethane (20 mL) was stirredat room temperature for 1 h and was then poured into ice water. Theorganic layer was separated and the aqueous phase was extracted withdichloromethane (50 mL×3). The combined extracts were washed with waterand aqueous NaHCO₃ solution, dried over Na₂SO₄ and evaporated underreduced pressure to afford4-chloro-6-(chloromethyl)benzo[d][1,3]dioxole, which was directly usedin the next step.

2-(7-Chlorobenzo[d][1,3]dioxol-5-yl)acetonitrile

A mixture of 4-chloro-6-(chloromethyl)benzo[d][1,3]dioxole (6.0 g, 29mmol) and NaCN (1.6 g, 32 mmol) in DMSO (20 mL) was stirred at 40° C.for 1 h and was then poured into water. The mixture was extracted withEtOAc (30 mL×3). The combined organic layers were washed with water andbrine, dried over Na₂SO₄ and evaporated under reduced pressure to afford2-(7-chlorobenzo[d][1,3]dioxol-5-yl)acetonitrile (3.4 g, 58%). ¹H NMR δ6.81 (s, 1H), 6.71 (s, 1H), 6.07 (s, 2H), 3.64 (s, 2H). ¹³C-NMR 6149.2,144.3, 124.4, 122.0, 117.4, 114.3, 107.0, 102.3, 23.1.

Example 19: 1-(Benzo[d]oxazol-5-yl)cyclopropanecarboxylic Acid

1-Benzooxazol-5-yl-cyclopropanecarboxylic Acid Methyl Ester

To a solution of 1-(3-amino-4-hydroxyphenyl)cyclopropanecarboxylic acidmethyl ester (3.00 g, 14.5 mmol) in DMF were added trimethylorthoformate (5.30 g, 14.5 mmol) and a catalytic amount ofp-tolueneslufonic acid monohydrate (0.3 g) at room temperature. Themixture was stirred for 3 hours at room temperature. The mixture wasdiluted with water and extracted with EtOAc (100 mL×3). The combinedorganic layers were dried over anhydrous Na₂SO₄ and evaporated undervacuum to give 1-benzooxazol-5-yl-cyclopropanecarboxylic acid methylester (3.1 g), which was directly used in the next step. ¹H NMR (CDCl₃,400 MHz) δ 8.09 (s, 1), 7.75 (d, J=1.2 Hz, 1H), 7.53-7.51 (m, 1H),7.42-7.40 (m, 1H), 3.66 (s, 3H), 1.69-1.67 (m, 2H), 1.27-1.24 (m, 2H).

1-(Benzo[d]oxazol-5-yl)cyclopropanecarboxylic Acid

To a solution of 1-benzooxazol-5-yl-cyclopropanecarboxylic acid methylester (2.9 g) in EtSH (30 mL) was added AlCl₃ (5.3 g, 40 mmol) inportions at 0° C. The reaction mixture was stirred for 18 hours at roomtemperature. Water (20 mL) was added dropwise at 0° C. The resultingmixture was extracted with EtOAc (100 mL×3). The combined organic layerswere dried over anhydrous Na₂SO₄ and evaporated under vacuum to give thecrude product, which was purified by column chromatography on silica gel(petroleum ether/ethyl acetate 1:2) to give1-(benzo[d]oxazol-5-yl)cyclopropanecarboxylic acid (280 mg, 11% over twosteps). ¹H NMR (DMSO, 400 MHz) δ 12.25 (brs, 1H), 8.71 (s, 1H),7.70-7.64 (m, 2H), 7.40 (dd, J=1.6, 8.4 Hz, 1H), 1.49-1.46 (m, 2H),1.21-1.18 (m, 2H). MS (ESI) m/e (M+H⁺) 204.4.

Example 20: 2-(7-Fluorobenzo[d][1,3]dioxol-5-yl)acetonitrile

3-Fluoro-4,5-dihydroxy-benzaldehyde

To a suspension of 3-fluoro-4-hydroxy-5-methoxy-benzaldehyde (1.35 g,7.94 mmol) in dichloromethane (100 mL) was added BBr₃ (1.5 mL, 16 mmol)dropwise at −78° C. under N₂. After addition, the mixture was warmed to−30° C. and it was stirred at this temperature for 5 h. The reactionmixture was poured into ice water. The precipitated solid was collectedby filtration and washed with dichloromethane to afford3-fluoro-4,5-dihydroxy-benzaldehyde (1.1 g, 89%), which was directlyused in the next step.

7-Fluoro-benzo[1,3]dioxole-5-carbaldehyde

To a solution of 3-fluoro-4,5-dihydroxy-benzaldehyde (1.5 g, 9.6 mmol)and BrClCH₂ (4.9 g, 38.5 mmol) in dry DMF (50 mL) was added Cs₂CO₃ (12.6g, 39 mmol). The mixture was stirred at 60° C. overnight and was thenpoured into water. The resulting mixture was extracted with EtOAc (50mL×3). The combined organic layers were washed with brine (100 mL),dried over Na₂SO₄ and evaporated under reduced pressure to give thecrude product, which was purified by column chromatography on silica gel(petroleum ether/ethyl acetate=10/1) to afford7-fluoro-benzo[1,3]dioxole-5-carbaldehyde (0.80 g, 49%). ¹H NMR (300MHz, CDCl₃) δ 9.78 (d, J=0.9 Hz, 1H), 7.26 (dd, J=1.5, 9.3 Hz, 1H), 7.19(d, J=1.2 Hz, 1H), 6.16 (s, 2H).

(7-Fluoro-benzo[1,3]dioxol-5-yl)-methanol

To a solution of 7-fluoro-benzo[1,3]dioxole-5-carbaldehyde (0.80 g, 4.7mmol) in MeOH (50 mL) was added NaBH₄ (0.36 g, 9.4 mmol) in portions at0° C. The mixture was stirred at this temperature for 30 min and wasthen concentrated to dryness. The residue was dissolved in EtOAc. TheEtOAc layer was washed with water, dried over Na₂SO₄ and concentrated todryness to afford (7-fluoro-benzo[1,3]dioxol-5-yl)-methanol (0.80 g,98%), which was directly used in the next step.

6-Chloromethyl-4-fluoro-benzo[1,3]dioxole

To SOCl₂ (20 mL) was added (7-fluoro-benzo[1,3]dioxol-5-yl)-methanol(0.80 g, 4.7 mmol) in portions at 0° C. The mixture was warmed to roomtemperature over 1 h and then was heated at reflux for 1 h. The excessSOCl₂ was evaporated under reduced pressure to give the crude product,which was basified with saturated aqueous NaHCO₃ to pH˜7. The aqueousphase was extracted with EtOAc (50 mL×3). The combined organic layerswere dried over Na₂SO₄ and evaporated under reduced pressure to give6-chloromethyl-4-fluoro-benzo[1,3]dioxole (0.80 g, 92%), which wasdirectly used in the next step.

2-(7-Fluorobenzo[d][1,3]dioxol-5-yl)acetonitrile

A mixture of 6-chloromethyl-4-fluoro-benzo[1,3]dioxole (0.80 g, 4.3mmol) and NaCN (417 mg, 8.51 mmol) in DMSO (20 mL) was stirred at 30° C.for 1 h and was then poured into water. The mixture was extracted withEtOAc (50 mL×3). The combined organic layers were washed with water (50mL) and brine (50 mL), dried over Na₂SO₄ and evaporated under reducedpressure to give the crude product, which was purified by columnchromatography on silica gel (petroleum ether/ethyl acetate=10/1) toafford 2-(7-fluorobenzo[d][1,3]dioxol-5-yl)acetonitrile (530 mg, 70%).¹H NMR (300 MHz, CDCl₃) δ 6.68-6.64 (m, 2H), 6.05 (s, 2H), 3.65 (s, 2H).¹³C-NMR 6151.1, 146.2, 134.1, 124.2, 117.5, 110.4, 104.8, 102.8, 23.3.

Example 21: 1-(1H-Indol-5-yl)cyclopropanecarboxylic Acid

Methyl 1-phenylcyclopropanecarboxylate

To a solution of 1-phenylcyclopropanecarboxylic acid (25 g, 0.15 mol) inCH₃OH (200 mL) was added TsOH (3 g, 0.1 mol) at room temperature. Themixture was refluxed overnight. The solvent was evaporated under reducedpressure to give crude product, which was dissolved into EtOAc. TheEtOAc layer was washed with aq. sat. NaHCO₃. The organic layer was driedover anhydrous Na₂SO₄ and evaporated under reduced pressure to givemethyl 1-phenylcyclopropanecarboxylate (26 g, 96%), which was useddirectly in the next step. ¹H NMR (400 MHz, CDCl₃) δ 7.37-7.26 (m, 5H),3.63 (s, 3H), 1.63-1.60 (m, 2H), 1.22-1.19 (m, 2H).

Methyl 1-(4-nitrophenyl)cyclopropanecarboxylate

To a solution of 1-phenylcyclopropanecarboxylate (20.62 g, 0.14 mol) inH₂SO₄/CH₂Cl₂ (40 mL/40 mL) was added KNO₃ (12.8 g, 0.13 mol) in portionat 0° C. The mixture was stirred for 0.5 hr at 0° C. Ice water was addedand the mixture was extracted with EtOAc (100 mL×3). The organic layerswere dried with anhydrous Na₂SO₄ and evaporated to give methyl1-(4-nitrophenyl)cyclopropanecarboxylate (21 g, 68%), which was useddirectly in the next step. ¹H NMR (300 MHz, CDCl₃) δ 8.18 (dd, J=2.1,6.9 Hz, 2H), 7.51 (dd, J=2.1, 6.9 Hz, 2H), 3.64 (s, 3H), 1.72-1.69 (m,2H), 1.25-1.22 (m, 2H).

Methyl 1-(4-aminophenyl)cyclopropanecarboxylate

To a solution of methyl 1-(4-nitrophenyl)cyclopropanecarboxylate (20 g,0.09 mol) in MeOH (400 mL) was added Ni (2 g) under nitrogen atmosphere.The mixture was stirred under hydrogen atmosphere (1 atm) at roomtemperature overnight. The catalyst was filtered off through a pad ofCelite and the filtrate was evaporated under vacuum to give crudeproduct, which was purified by chromatography column on silica gel(petroleum ether/ethyl acetate=10:1) to give methyl1-(4-aminophenyl)cyclopropanecarboxylate (11.38 g, 66%). ¹H NMR (300MHz, CDCl₃) δ 7.16 (d, J=8.1 Hz, 2H), 6.86 (d, J=7.8 Hz, 2H), 4.31 (br,2H), 3.61 (s, 3H), 1.55-1.50 (m, 2H), 1.30-1.12 (m, 2H).

Methyl 1-(4-amino-3-bromophenyl)cyclopropanecarboxylate

To a solution of methyl 1-(4-aminophenyl)cyclopropanecarboxylate (10.38g, 0.05 mol) in acetonitrile (200 mL) was added NBS (9.3 g, 0.05 mol) atroom temperature. The mixture was stirred overnight. Water (200 mL) wasadded. The organic layer was separated and the aqueous phase wasextracted with EtOAc (80 mL×3). The organic layers were dried withanhydrous Na₂SO₄ and evaporated to give methyl1-(4-amino-3-bromophenyl)cyclopropanecarboxylate (10.6 g, 78%), whichwas used directly in the next step. ¹H NMR (400 MHz, CDCl₃) δ 7.38 (d,J=2.0 Hz, 1H), 7.08 (dd, J=1.6, 8.4 Hz, 1H), 6.70 (d, J=8.4 Hz, 1H),3.62 (s, 3H), 1.56-1.54 (m, 2H), 1.14-1.11 (m, 2H).

Methyl 1-(4-amino-3-((trimethylsilyl)ethynyl)phenyl)cyclopropaneCarboxylate

To a degassed solution of methyl 1-(4-amino-3-bromophenyl)cyclopropanecarboxylate (8 g, 0.03 mol) in Et₃N (100 mL) was addedethynyl-trimethyl-silane (30 g, 0.3 mol), DMAP (5% mol) and Pd(PPh₃)₂Cl₂(5% mol) under N₂. The mixture was refluxed at 70° C. overnight. Theinsoluble solid was filtered off and washed with EtOAc (100 mL×3). Thefiltrate was evaporated under reduced pressure to give a residue, whichwas purified by chromatography column on silica gel (petroleumether/ethyl acetate=20:1) to give methyl1-(4-amino-3-((trimethylsilyl)ethynyl)phenyl)cyclopropanecarboxylate(4.8 g, 56%). ¹H NMR (300 MHz, CDCl₃) δ 7.27 (s, 1H), 7.10 (dd, J=2.1,8.4 Hz, 1H), 6.64 (d, J=8.4 Hz, 1H), 3.60 (s, 3H), 1.55-1.51 (m, 2H),1.12-1.09 (m, 2H), 0.24 (s, 9H).

Methyl 1-(1H-indol-5-yl)cyclopropanecarboxylate

To a degassed solution of methyl1-(4-amino-3-((trimethylsilyl)ethynyl)phenyl) cyclopropanecarboxylate(4.69 g, 0.02 mol) in DMF (20 mL) was added CuI (1.5 g, 0.008 mol) underN₂ at room temperature. The mixture was stirred for 3 hr at roomtemperature. The insoluble solid was filtered off and washed with EtOAc(50 mL×3). The filtrate was evaporated under reduced pressure to give aresidue, which was purified by chromatography column on silica gel(petroleum ether/ethyl acetate=20:1) to give methyl1-(1H-indol-5-yl)cyclopropanecarboxylate (2.2 g, 51%). ¹H NMR (400 MHz,CDCl₃) δ 7.61 (s, 1H), 7.33 (d, J=8.4 Hz, 1H), 7.23-7.18 (m, 2H),6.52-6.51 (m, 1H) 3.62 (s, 3H), 1.65-1.62 (m, 2H), 1.29-1.23 (m, 2H)

1-(1H-Indol-5-yl)cyclopropanecarboxylic Acid

To a solution of methyl 1-(1H-indol-5-yl)cyclopropanecarboxylate (1.74g, 8 mmol) in CH₃OH (50 m L) and water (20 mL) was added LiOH (1.7 g,0.04 mol). The mixture was heated at 45° C. for 3 hr. Water was addedand the mixture was acidified with concentrated HCl to pH˜3 before beingextracted with EtOAc (20 mL×3). The organic layers were dried overanhydrous Na₂SO₄ and evaporated to give1-(1H-indol-5-yl)cyclopropanecarboxylic acid (1.4 g, 87%). ¹H NMR (300MHz, DMSO-d₆) 7.43 (s, 1H), 7.30-7.26 (m, 2H), 7.04 (dd, J=1.5, 8.4 Hz,1H), 6.35 (s, 1H), 1.45-1.41 (m, 2H), 1.14-1.10 (m, 2H).

Example 22: 1-(4-Oxochroman-6-yl)cyclopropanecarboxylic Acid

1-[4-(2-tert-Butoxycarbonyl-ethoxy)-phenyl]-cyclopropanecarboxylicMethyl Ester

To a solution of 1-(4-hydroxy-phenyl)-cyclopropanecarboxylic methylester (7.0 g, 3.6 mmol) in acrylic tert-butyl ester (50 mL) was added Na(42 mg, 1.8 mmol) at room temperature. The mixture was heated at 110° C.for 1 h. After cooling to room temperature, the resulting mixture wasquenched with water and extracted with EtOAc (100 mL×3). The combinedorganic extracts were dried over anhydrous Na₂SO₄ and evaporated undervacuum to give the crude product, which was purified by columnchromatography on silica gel (petroleum ether/ethyl acetate 20:1) togive 1-[4-(2-tert-butoxycarbonyl-ethoxy)-phenyl]-cyclopropanecarboxylicmethyl ester (6.3 g, 54%) and unreacted start material (3.0 g). ¹H NMR(300 MHz, CDCl₃) δ 7.24 (d, J=8.7 Hz, 2H), 6.84 (d, J=8.7 Hz, 2H), 4.20(t, J=6.6 Hz, 2H), 3.62 (s, 3H), 2.69 (t, J=6.6 Hz, 2H), 1.59-1.56 (m,2H), 1.47 (s, 9H), 1.17-0.42 (m, 2H).

1-[4-(2-Carboxy-ethoxy)-phenyl]-cyclopropanecarboxylic Methyl Ester

A solution of1-[4-(2-tert-butoxycarbonyl-ethoxy)-phenyl]-cyclopropanecarboxylicmethyl ester (6.3 g, 20 mmol) in HCl (20%, 200 mL) was heated at 110° C.for 1 h. After cooling to room temperature, the resulting mixture wasfiltered. The solid was washed with water and dried under vacuum to give1-[4-(2-carboxy-ethoxy)-phenyl]-cyclopropanecarboxylic methyl ester (5.0g, 96%). ¹H NMR (300 MHz, DMSO) δ 7.23-7.19 (m, 2H), 6.85-6.81 (m, 2H),4.13 (t, J=6.0 Hz, 2H), 3.51 (s, 3H), 2.66 (t, J=6.0 Hz, 2H), 1.43-1.39(m, 2H), 1.14-1.10 (m, 2H).

1-(4-Oxochroman-6-yl)cyclopropanecarboxylic Acid

To a solution of 1-[4-(2-carboxy-ethoxy)-phenyl]-cyclopropanecarboxylicmethyl ester (5.0 g, 20 mmol) in CH₂Cl₂ (50 mL) were added oxalylchloride (4.8 g, 38 mmol) and two drops of DMF at 0° C. The mixture wasstirred at 0-5° C. for 1 h and then evaporated under vacuum. To theresulting mixture was added CH₂Cl₂ (50 mL) at 0° C. and stirring wascontinued at 0-5° C. for 1 h. The reaction was slowly quenched withwater and was extracted with EtOAc (50 mL×3). The combined organicextracts were dried over anhydrous Na₂SO₄ and evaporated under vacuum togive the crude product, which was purified by column chromatography onsilica gel (petroleum ether/ethyl acetate 20:1-2:1) to give1-(4-oxochroman-6-yl)cyclopropanecarboxylic acid (830 mg, 19%) andmethyl 1-(4-oxochroman-6-yl)cyclopropanecarboxylate (1.8 g, 38%).1-(4-Oxochroman-6-yl)cyclopropane-carboxylic acid: ¹H NMR (400 MHz,DMSO) δ 12.33 (br s, 1H), 7.62 (d, J=2.0 Hz, 1H), 7.50 (dd, J=2.4, 8.4Hz, 1H), 6.95 (d, J=8.4 Hz, 1H), 4.50 (t, J=6.4 Hz, 2H), 2.75 (t, J=6.4Hz, 2H), 1.44-1.38 (m, 2H), 1.10-1.07 (m, 2H). MS (ESI) m/z (M+H⁺)231.4. 1-(4-Oxochroman-6-yl)cyclopropanecarboxylate: ¹H NMR (400 MHz,CDCl₃) δ 7.83 (d, J=2.4 Hz, 1H), 7.48 (dd, J=2.4, 8.4 Hz, 1H), 6.93 (d,J=8.4 Hz, 1H), 4.55-4.52 (m, 2H), 3.62 (s, 3H), 2.80 (t, J=6.4 Hz, 2H),1.62-1.56 (m, 2H), 1.18-1.15 (m, 2H).

Example 23: 1-(4-Hydroxy-4-methoxychroman-6-yl)cyclopropanecarboxylicAcid

1-(4-Hydroxy-4-methoxychroman-6-yl)cyclopropanecarboxylic Acid

To a solution of methyl 1-(4-oxochroman-6-yl)cyclopropanecarboxylate(1.0 g, 4.1 mmol) in MeOH (20 mL) and water (20 mL) was added LiOH H₂O(0.70 g, 16 mmol) in portions at room temperature. The mixture wasstirred overnight at room temperature before the MeOH was removed byevaporation under vacuum. Water and Et₂O were added to the residue andthe aqueous layer was separated, acidified with HCl and extracted withEtOAc (50 mL×3). The combined organic extracts dried over anhydrousNa₂SO₄ and evaporated under vacuum to give1-(4-hydroxy-4-methoxychroman-6-yl)cyclopropanecarboxylic acid (480 mg,44%). ¹H NMR (400 MHz, CDCl₃) δ 12.16 (s, 1H), 7.73 (d, J=2.0 Hz, 1H),7.47 (dd, J=2.0, 8.4 Hz, 1H), 6.93 (d, J=8.8 Hz, 1H), 3.83-3.80 (m, 2H),3.39 (s, 3H), 3.28-3.25 (m, 2H), 1.71-1.68 (m, 2H), 1.25-1.22 (m, 2H).MS (ESI) m/z (M+H⁺) 263.1.

Example 24: 1-(4-Hydroxy-4-methoxychroman-6-yl)cyclopropanecarboxylicAcid

1-Chroman-6-yl-cyclopropanecarboxylic Methyl Ester

To trifluoroacetic acid (20 mL) was added NaBH₄ (0.70 g, 130 mmol) inportions at 0° C. under N₂ atmosphere. After stirring for 5 min, asolution of 1-(4-oxo-chroman-6-yl)-cyclopropanecarboxylic methyl ester(1.6 g, 6.5 mmol) was added at 15° C. The reaction mixture was stirredfor 1 h at room temperature before being slowly quenched with water. Theresulting mixture was extracted with EtOAc (50 mL×3). The combinedorganic extracts dried over anhydrous Na₂SO₄ and evaporated under vacuumto give 1-chroman-6-yl-cyclopropanecarboxylic methyl ester (1.4 g, 92%),which was used directly in the next step. ¹H NMR (300 MHz, CDCl₃) δ7.07-7.00 (m, 2H), 6.73 (d, J=8.4 Hz, 1H), 4.17 (t, J=5.1 Hz, 2H), 3.62(s, 3H), 2.79-2.75 (m, 2H), 2.05-1.96 (m, 2H), 1.57-1.54 (m, 2H),1.16-1.13 (m, 2H).

1-(4-Hydroxy-4-methoxychroman-6-yl)cyclopropanecarboxylic Acid

To a solution of 1-chroman-6-yl-cyclopropanecarboxylic methyl ester (1.4g, 60 mmol) in MeOH (20 mL) and water (20 mL) was added LiOH H₂O (1.0 g,240 mmol) in portions at room temperature. The mixture was stirredovernight at room temperature before the MeOH was removed by evaporationunder vacuum. Water and Et₂O were added and the aqueous layer wasseparated, acidified with HCl and extracted with EtOAc (50 mL×3). Thecombined organic extracts dried over anhydrous Na₂SO₄ and evaporatedunder vacuum to give1-(4-Hydroxy-4-methoxychroman-6-yl)cyclopropanecarboxylic acid (1.0 g,76%). ¹H NMR (400 MHz, DMSO) δ 12.10 (br s, 1H), 6.95 (d, J=2.4 Hz, 2H),6.61-6.59 (m, 1H), 4.09-4.06 (m, 2H), 2.70-2.67 (m, 2H), 1.88-1.86 (m,2H), 1.37-1.35 (m, 2H), 1.04-1.01 (m, 2H). MS (ESI) m/z (M+H⁺) 217.4.

Example 25: 1-(3-Methylbenzo[d]isoxazol-5-yl)cyclopropanecarboxylic Acid

1-(3-Acetyl-4-hydroxy-phenyl)-cyclopropanecarboxylic Methyl Ester

To a stirred suspension of AlCl₃ (58 g, 440 mmol) in CS₂ (500 mL) wasadded acetyl chloride (7.4 g, 95 mmol) at room temperature. Afterstirring for 5 min, methyl 1-(4-methoxyphenyl)cyclopropanecarboxylate(15 g, 73 mmol) was added. The reaction mixture was heated at reflux for2 h before ice water was added carefully to the mixture at roomtemperature. The resulting mixture was extracted with EtOAc (150 mL×3).The combined organic extracts were dried over anhydrous Na₂SO₄ andevaporated under reduced pressure to give1-(3-acetyl-4-hydroxy-phenyl)-cyclopropanecarboxylic methyl ester (15 g,81%), which was used in the next step without further purification. ¹HNMR (CDCl₃, 400 MHz) δ 12.28 (s, 1H), 7.67 (d, J=2.0 Hz, 1H), 7.47 (dd,J=2.0, 8.4 Hz, 1H), 6.94 (d, J=8.4 Hz, 1H), 3.64 (s, 3H), 2.64 (s, 3H),1.65-1.62 (m, 2H), 1.18-1.16 (m, 2H).

1-[4-Hydroxy-3-(1-hydroxyimino-ethyl)-phenyl]-cyclopropanecarboxylicMethyl Ester

To a stirred solution of1-(3-acetyl-4-hydroxy-phenyl)-cyclopropanecarboxylic methyl ester (14.6g, 58.8 mmol) in EtOH (500 mL) were added hydroxylamine hydrochloride(9.00 g, 129 mmol) and sodium acetate (11.6 g, 141 mmol) at roomtemperature. The resulting mixture was heated at reflux overnight. Afterremoval of EtOH under vacuum, water (200 mL) and EtOAc (200 mL) wereadded. The organic layer was separated and the aqueous layer wasextracted with EtOAc (100 mL×3). The combined organic layers were driedover anhydrous Na₂SO₄ and evaporated under vacuum to give1-[4-hydroxy-3-(1-hydroxyimino-ethyl)-phenyl]-cyclopropanecarboxylicmethyl ester (14.5 g, 98%), which was used in the next step withoutfurther purification. ¹H NMR (CDCl₃, 400 MHz) δ 11.09 (s, 1H), 7.39 (d,J=2.0 Hz, 1H), 7.23 (d, J=2.0 Hz, 1H), 7.14 (s, 1H), 6.91 (d, J=8.4 Hz,1H), 3.63 (s, 3H), 2.36 (s, 3H), 1.62-1.59 (m, 2H), 1.18-1.15 (m, 2H).

(E)-Methyl 1-(3-(1-(acetoxyimino)ethyl)-4-hydroxyphenyl)cyclopropaneCarboxylate

The solution of1-[4-hydroxy-3-(1-hydroxyimino-ethyl)-phenyl]-cyclopropanecarboxylicmethyl ester (10.0 g, 40.1 mmol) in Ac₂O (250 mL) was heated at 45° C.for 4 h. The Ac₂O was removed by evaporation under vacuum before water(100 mL) and EtOAc (100 mL) were added. The organic layer was separatedand the aqueous layer was extracted with EtOAc (100 mL×2). The combinedorganic layers were dried over anhydrous Na₂SO₄ and evaporated undervacuum to give (E)-methyl1-(3-(1-(acetoxyimino)ethyl)-4-hydroxyphenyl)cyclopropanecarboxylate(10.5 g, 99%), which was used in the next step without furtherpurification.

Methyl 1-(3-methylbenzo[d]isoxazol-5-yl)cyclopropanecarboxylate

A solution of (E)-methyl1-(3-(1-(acetoxyimino)ethyl)-4-hydroxyphenyl)cyclopropane carboxylate(10.5 g, 39.6 mmol) and pyridine (31.3 g, 396 mmol) in DMF (150 mL) washeated at 125° C. for 10 h. The cooled reaction mixture was poured intowater (250 mL) and was extracted with EtOAc (100 mL×3). The combinedorganic layers were dried over anhydrous Na₂SO₄ and evaporated undervacuum to give the crude product, which was purified by columnchromatography on silica gel (petroleum ether/ethyl acetate 50:1) togive methyl 1-(3-methylbenzo[d]isoxazol-5-yl)cyclopropanecarboxylate(7.5 g, 82%). ¹H NMR (CDCl₃ 300 MHz) δ 7.58-7.54 (m, 2H), 7.48 (dd,J=1.5, 8.1 Hz, 1H), 3.63 (s, 3H), 2.58 (s, 3H), 1.71-1.68 (m, 2H),1.27-1.23 (m, 2H).

1-(3-Methylbenzo[d]isoxazol-5-yl)cyclopropanecarboxylic Acid

To a solution of methyl1-(3-methylbenzo[d]isoxazol-5-yl)cyclopropanecarboxylate (1.5 g, 6.5mmol) in MeOH (20 mL) and water (2 mL) was added LiOH H₂O (0.80 g, 19mmol) in portions at room temperature. The reaction mixture was stirredat room temperature overnight before the MeOH was removed by evaporationunder vacuum. Water and Et₂O were added and the aqueous layer wasseparated, acidified with HCl and extracted with EtOAc (50 mL×3). Thecombined organic extracts were dried over anhydrous Na₂SO₄ andevaporated under vacuum to give1-(3-methylbenzo[d]isoxazol-5-yl)cyclopropanecarboxylic acid (455 mg,32%). ¹H NMR (400 MHz, DMSO) δ 12.40 (br s, 1H), 7.76 (s, 1H), 7.60-7.57(m, 2H), 2.63 (s, 3H), 1.52-1.48 (m, 2H), 1.23-1.19 (m, 2H). MS (ESI)m/z (M+H⁺) 218.1.

Example 26:1-(Spiro[benzo[d][1,3]dioxole-2,1′-cyclobutane]-5-yl)cyclopropaneCarboxylic Acid

1-(3,4-Dihydroxy-phenyl)-cyclopropanecarboxylic Methyl Ester

To a solution of 1-(3,4-dihydroxyphenyl)cyclopropanecarboxylic acid (4.5g) in MeOH (30 mL) was added TsOH (0.25 g, 1.3 mmol). The stirring wascontinued at 50° C. overnight before the mixture was cooled to roomtemperature. The mixture was concentrated under vacuum and the residuewas purified by column chromatography on silica gel (petroleumether/ethyl acetate 3:1) to give1-(3,4-dihydroxy-phenyl)-cyclopropanecarboxylic methyl ester (2.1 g). ¹HNMR (DMSO 300 MHz) δ 8.81 (brs, 2H), 6.66 (d, J=2.1 Hz, 1H), 6.61 (d,J=8.1 Hz, 1H), 6.53 (dd, J=2.1, 8.1 Hz, 1H), 3.51 (s, 3H), 1.38-1.35 (m,2H), 1.07-1.03 (m, 2H).

Methyl 1-(spiro[benzo[d][1,3]dioxole-2,1′-cyclobutane]-5-yl)cyclopropaneCarboxylate

To a solution of 1-(3,4-dihydroxy-phenyl)-cyclopropanecarboxylic methylester (1.0 g, 4.8 mmol) in toluene (30 mL) was added TsOH (0.10 g, 0.50mmol) and cyclobutanone (0.70 g, 10 mmol). The reaction mixture washeated at reflux for 2 h before being concentrated under vacuum. Theresidue was purified by chromatography on silica gel (petroleumether/ethyl acetate 15:1) to give methyl1-(spiro[benzo[d][1,3]dioxole-2,1′-cyclobutane]-5-yl)cyclopropanecarboxylate(0.6 g, 50%). ¹H NMR (CDCl₃ 300 MHz) δ 6.78-6.65 (m, 3H), 3.62 (s, 3H),2.64-2.58 (m, 4H), 1.89-1.78 (m, 2H), 1.56-1.54 (m, 2H), 1.53-1.12 m,2H).

1-(Spiro[benzo[d][1,3]dioxole-2,1′-cyclobutane]-5-yl)cyclopropanecarboxylic Acid

To a mixture of methyl1-(spiro[benzo[d][1,3]dioxole-2,1′-cyclobutane]-5-yl)cycl-opropanecarboxylate(0.60 g, 2.3 mmol) in THF/H₂O (4:1, 10 mL) was added LiOH (0.30 g, 6.9mmol). The mixture was stirred at 60° C. for 24 h. HCl (0.5 N) was addedslowly to the mixture at 0° C. until pH 2-3. The mixture was extractedwith EtOAc (10 mL×3). The combined organic phases were washed withbrine, dried over anhydrous MgSO₄, and washed with petroleum ether togive 1-(spiro[benzo[d][1,3]-dioxole-2,1′-cyclobutane]-5-yl)cyclopropanecarboxylic acid (330 mg, 59%). ¹HNMR (400 MHz, CDCl₃) δ 6.78-6.65 (m,3H), 2.65-2.58 (m, 4H), 1.86-1.78 (m, 2H), 1.63-1.60 (m, 2H), 1.26-1.19(m, 2H).

Example 27: 2-(2,3-Dihydrobenzo[b][1,4]dioxin-6-yl)acetonitrile

2,3-Dihydro-benzo[1,4]dioxine-6-carboxylic Acid Ethyl Ester

To a suspension of Cs₂CO₃ (270 g, 1.49 mol) in DMF (1000 mL) were added3,4-dihydroxybenzoic acid ethyl ester (54.6 g, 0.3 mol) and1,2-dibromoethane (54.3 g, 0.29 mol) at room temperature. The resultingmixture was stirred at 80° C. overnight and then poured into ice-water.The mixture was extracted with EtOAc (200 mL×3). The combined organiclayers were washed with water (200 mL×3) and brine (100 mL), dried overNa₂SO₄ and concentrated to dryness. The residue was purified by column(petroleum ether/ethyl acetate 50:1) on silica gel to obtain2,3-dihydro-benzo[1,4]dioxine-6-carboxylic acid ethyl ester (18 g, 29%).¹H NMR (300 MHz, CDCl₃) δ 7.53 (dd, J=1.8, 7.2 Hz, 2H), 6.84-6.87 (m,1H), 4.22-4.34 (m, 6H), 1.35 (t, J=7.2 Hz, 3H).

(2,3-Dihydro-benzo[1,4]dioxin-6-yl)-methanol

To a suspension of LiAlH₄ (2.8 g, 74 mmol) in THF (20 mL) was addeddropwise a solution of 2,3-dihydro-benzo[1,4]dioxine-6-carboxylic acidethyl ester (15 g, 72 mmol) in THF (10 mL) at 0° C. under N₂. Themixture was stirred at room temperature for 1 h and then quenchedcarefully with addition of water (2.8 mL) and NaOH (10%, 28 mL) withcooling. The precipitated solid was filtered off and the filtrate wasevaporated to dryness to obtain(2,3-dihydro-benzo[1,4]dioxin-6-yl)-methanol (10.6 g). ¹H NMR (300 MHz,DMSO-d₆) δ 6.73-6.78 (m, 3H), 5.02 (t, J=5.7 Hz, 1H), 4.34 (d, J=6.0 Hz,2H), 4.17-4.20 (m, 4H).

6-Chloromethyl-2,3-dihydro-benzo[1,4]dioxine

A mixture of (2,3-dihydro-benzo[1,4]dioxin-6-yl)methanol (10.6 g) inSOCl₂ (10 mL) was stirred at room temperature for 10 min and then pouredinto ice-water. The organic layer was separated and the aqueous phasewas extracted with dichloromethane (50 mL×3). The combined organiclayers were washed with NaHCO₃ (sat solution), water and brine, driedover Na₂SO₄ and concentrated to dryness to obtain6-chloromethyl-2,3-dihydro-benzo[1,4]dioxine (12 g, 88% over two steps),which was used directly in next step.

2-(2,3-Dihydrobenzo[b][1,4]dioxin-6-yl)acetonitrile

A mixture of 6-chloromethyl-2,3-dihydro-benzo[1,4]dioxine (12.5 g, 67.7mmol) and NaCN (4.30 g, 87.8 mmol) in DMSO (50 mL) was stirred at rt for1 h. The mixture was poured into water (150 mL) and then extracted withdichloromethane (50 mL×4). The combined organic layers were washed withwater (50 mL×2) and brine (50 mL), dried over Na₂SO₄ and concentrated todryness. The residue was purified by column (petroleum ether/ethylacetate 50:1) on silica gel to obtain2-(2,3-dihydrobenzo[b][1,4]dioxin-6-yl)acetonitrile as a yellow oil(10.2 g, 86%). ¹H-NMR (300 MHz, CDCl₃) δ 6.78-6.86 (m, 3H), 4.25 (s,4H), 3.63 (s, 2H).

The following Table 2 contains a list of carboxylic acid building blocksthat were commercially available, or prepared by one of the threemethods described above:

TABLE 2 Carboxylic acid building blocks. Name Structure1-benzo[1,3]dioxol-5- ylcyclopropane-1-carboxylic acid

1-(2,2-difluorobenzo[1,3]dioxol-5- yl)cyclopropane-1-carboxylic acid

1-(3,4-dihydroxyphenyl)cyclopropanecarboxylic acid

1-(3-methoxyphenyl)cyclopropane-1-carboxylic acid

1-(2-methoxyphenyl)cyclopropane-1-carboxylic acid

1-[4-(trifluoromethoxy)phenyl]cyclopropane- 1-carboxylic acid

1-(2,2-dimethylbenzo[d][1,3]dioxol-5- yl)cyclopropanecarboxylic acid

tetrahydro-4-(4-methoxyphenyl)- 2H-pyran-4-carboxylic acid

1-phenylcyclopropane-1-carboxylic acid

1-(4-methoxyphenyl)cyclopropane-1-carboxylic acid

1-(4-chlorophenyl)cyclopropane-1-carboxylic acid

1-(3-hydroxyphenyl)cyclopropanecarboxylic acid

1-phenylcyclopentanecarboxylic acid

1-(2-oxo-2,3-dihydrobenzo[d]oxazol-5- yl)cyclopropanecarboxylic acid

1-(benzofuran-5-yl)cyclopropanecarboxylic acid

1-(4-methoxyphenyl)cyclohexanecarboxylic acid

1-(4-chlorophenyl)cyclohexanecarboxylic acid

1-(2,3-dihydrobenzofuran-5- yl)cyclopropanecarboxylic acid

1-(3,3-dimethyl-2,3-dihydrobenzofuran- 5-yl)cyclopropanecarboxylic acid

1-(7-methoxybenzo[d][1,3]dioxol-5- yl)cyclopropanecarboxylic acid

1-(3-hydroxy-4-methoxyphenyl) cyclopropanecarboxylic acid

1-(4-chloro-3-hydroxyphenyl) cyclopropanecarboxylic acid

1-(3-(benzyloxy)-4- chlorophenyl)cyclopropanecarboxylic acid

1-(4-chlorophenyl)cyclopentanecarboxylic acid

1-(3-(benzyloxy)-4- methoxyphenyl)cyclopropanecarboxylic acid

1-(3-chloro-4-methoxyphenyl) cyclopropanecarboxylic acid

1-(3-fluoro-4-methoxyphenyl) cyclopropanecarboxylic acid

1-(4-methoxy-3-methylphenyl) cyclopropanecarboxylic acid

1-(4-(benzyloxy)-3- methoxyphenyl)cyclopropanecarboxylic acid

1-(4-chloro-3-methoxyphenyl) cyclopropanecarboxylic acid

1-(3-chloro-4-hydroxyphenyl) cyclopropanecarboxylic acid

1-(3-(hydroxymethyl)-4- methoxyphenyl)cyclopropanecarboxylic acid

1-(4-methoxyphenyl)cyclopentanecarboxylic acid

1-phenylcyclohexanecarboxylic acid

1-(3,4-dimethoxyphenyl)cyclopropanecarboxylic acid

1-(7-chlorobenzo[d][1,3]dioxol-5- yl)cyclopropanecarboxylic acid

1-(benzo[d]oxazol-5-yl)cyclopropanecarboxylic acid

1-(7-fluorobenzo[d][1,3]dioxol-5- yl)cyclopropanecarboxylic acid

1-(3,4-difluorophenyl)cyclopropanecarboxylic acid

1-(1H-indol-5-yl)cyclopropanecarboxylic acid

1-(1H-benzo[d]imidazol-5- yl)cyclopropanecarboxylic acid

1-(2-methyl-1H-benzo[d]imidazol-5- yl)cyclopropanecarboxylic acid

1-(1-methyl-1H-benzo[d]imidazol- 5-yl)cyclopropanecarboxylic acid

1-(3-methylbenzo[d]isoxazol-5- yl)cyclopropanecarboxylic acid

1-(spiro[benzo[d][1,3]dioxole-2,1′-cyclobutane]-5-yl)cyclopropanecarboxylic acid

1-(1H-benzo[d][1,2,3]triazol-5- yl)cyclopropanecarboxylic acid

1-(1-methyl-1H-benzo[d][1,2,3]triazol- 5-yl)cyclopropanecarboxylic acid

1-(1,3-dihydroisobenzofuran-5- yl)cyclopropanecarboxylic acid

1-(6-fluorobenzo[d][1,3]dioxol-5- yl)cyclopropanecarboxylic acid

1-(2,3-dihydrobenzofuran-6- yl)cyclopropanecarboxylic acid

1-(chroman-6-yl)cyclopropanecarboxylic acid

1-(4-hydroxy-4-methoxychroman-6- yl)cyclopropanecarboxylic acid

1-(4-oxochroman-6-yl)cyclopropanecarboxylic acid

1-(3,4-dichlorophenyl)cyclopropanecarboxylic acid

1-(2,3-dihydrobenzo[b][1,4]dioxin-6- yl)cyclopropanecarboxylic acid

1-(benzofuran-6-yl)cyclopropanecarboxylic acid

Specific Procedures: Synthesis of Aminoindole Building Blocks

Example 28: 3-Methyl-1H-indol-6-amine

(3-Nitro-phenyl)-hydrazine Hydrochloride Salt

3-Nitro-phenylamine (27.6 g, 0.2 mol) was dissolved in the mixture ofH₂O (40 mL) and 37% HCl (40 mL). A solution of NaNO₂ (13.8 g, 0.2 mol)in H₂O (60 mL) was added to the mixture at 0° C., and then a solution ofSnCl₂.H₂O (135.5 g, 0.6 mol) in 37% HCl (100 mL) was added at thattemperature. After stirring at 0° C. for 0.5 h, the insoluble materialwas isolated by filtration and was washed with water to give(3-nitrophenyl)hydrazine hydrochloride (27.6 g, 730%).

N-(3-Nitro-phenyl)-N-propylidene-hydrazine

Sodium hydroxide solution (10%, 15 mL) was added slowly to a stirredsuspension of (3-nitrophenyl)hydrazine hydrochloride (1.89 g, 10 mmol)in ethanol (20 mL) until pH 6. Acetic acid (5 mL) was added to themixture followed by propionaldehyde (0.7 g, 12 mmol). After stirring for3 h at room temperature, the mixture was poured into ice-water and theresulting precipitate was isolated by filtration, washed with water anddried in air to obtain (E)-1-(3-nitrophenyl)-2-propylidenehydrazine,which was used directly in the next step.

3-Methyl-4-nitro-1H-indole 3 and 3-methyl-6-nitro-1H-indole

A mixture of (E)-1-(3-nitrophenyl)-2-propylidenehydrazine dissolved in85% H₃PO₄ (20 mL) and toluene (20 mL) was heated at 90-100° C. for 2 h.After cooling, toluene was removed under reduced pressure. The resultantoil was basified to pH 8 with 10% NaOH. The aqueous layer was extractedwith EtOAc (100 mL×3). The combined organic layers were dried, filteredand concentrated under reduced pressure to afford the mixture of3-methyl-4-nitro-1H-indole and 3-methyl-6-nitro-1H-indole [1.5 g intotal, 86%, two steps from (3-nitrophenyl)hydrazine hydrochloride] whichwas used to the next step without further purification.

3-Methyl-1H-indol-6-amine

The crude mixture from previous steps (3 g, 17 mmol) and 10% Pd-C (0.5g) in ethanol (30 mL) was stirred overnight under H₂ (1 atm) at roomtemperature. Pd-C was filtered off and the filtrate was concentratedunder reduced pressure. The solid residue was purified by column to give3-methyl-1H-indol-6-amine (0.6 g, 24%). ¹H NMR (CDCl₃) δ 7.59 (br s.1H), 7.34 (d, J=8.0 Hz, 1H), 6.77 (s, 1H), 6.64 (s, 1H), 6.57 (m, 1H),3.57 (brs, 2H), 2.28 (s, 3H); MS (ESI) m/e (M+H⁺) 147.2.

Example 29: 3-tert-Butyl-1H-indol-5-amine

3-tert-Butyl-5-nitro-1H-indole

To a mixture of 5-nitro-1H-indole (6.0 g, 37 mmol) and AlCl₃ (24 g, 0.18mol) in CH₂Cl₂ (100 mL) at 0° C. was added 2-bromo-2-methyl-propane (8.1g, 37 mmol) dropwise. After being stirred at 15° C. overnight, themixture was poured into ice (100 mL). The precipitated salts wereremoved by filtration and the aqueous layer was extracted with CH₂Cl₂(30 mL×3). The combined organic layers were washed with water, brine,dried over Na₂SO₄ and concentrated under vacuum to obtain the crudeproduct, which was purified by column chromatography on silica gel(petroleum ether/ethyl acetate=20:1) to give3-tert-butyl-5-nitro-1H-indole (2.5 g, 31%). ¹H NMR (CDCl₃, 400 MHz) δ8.49 (d, J=1.6 Hz, 1H), 8.31 (brs, 1H), 8.05 (dd, J=2.0, 8.8 Hz, 1H),7.33 (d, J=8.8 Hz, 1H), 6.42 (d, J=1.6 Hz, 1H), 1.42 (s, 9H).

3-tert-Butyl-1H-indol-5-amine

To a solution of 3-tert-butyl-5-nitro-1H-indole (2.5 g, 12 mmol) in MeOH(30 mL) was added Raney Nickel (0.2 g) under N₂ protection. The mixturewas stirred under hydrogen atmosphere (1 atm) at 15° C. for 1 h. Thecatalyst was filtered off and the filtrate was concentrated to drynessunder vacuum. The residue was purified by preparative HLPC to afford3-tert-butyl-1H-indol-5-amine (0.43 g, 19%). ¹H NMR (CDCl₃, 400 MHz) δ7.72 (br.s, 1H), 7.11 (d, J=8.4 Hz, 1H), 6.86 (d, J=2.0 Hz, 1H), 6.59(dd, J=2.0, 8.4 Hz, 1H), 6.09 (d, J=1.6 Hz, 1H), 1.37 (s, 9H); MS (ESI)m/e (M+H⁺) 189.1.

Example 30: 2-tert-Butyl-6-fluoro-1H-indol-5-amine and6-tert-butoxy-2-tert-butyl-1H-indol-5-amine

2-Bromo-5-fluoro-4-nitroaniline

To a mixture of 3-fluoro-4-nitroaniline (6.5 g, 42.2 mmol) in AcOH (80mL) and chloroform (25 mL) was added dropwise Br₂ (2.15 mL, 42.2 mmol)at 0° C. After addition, the resulting mixture was stirred at roomtemperature for 2 h and then poured into ice water. The mixture wasbasified with aqueous NaOH (10%) to pH˜8.0-9.0 under cooling and thenextracted with EtOAc (50 mL×3). The combined organic layers were washedwith water (80 mL×2) and brine (100 mL), dried over Na₂SO₄ andconcentrated under reduced pressure to give2-bromo-5-fluoro-4-nitroaniline (9 g, 90%). ¹H-NMR (400 MHz, DMSO-d₆) δ8.26 (d, J=8.0, Hz, 1H), 7.07 (brs, 2H), 6.62 (d, J=9.6 Hz, 1H).

2-(3,3-Dimethylbut-1-ynyl)-5-fluoro-4-nitroaniline

A mixture of 2-bromo-5-fluoro-4-nitroaniline (9.0 g, 38.4 mmol),3,3-dimethyl-but-1-yne (9.95 g, 121 mmol), CuI (0.5 g 2.6 mmol),Pd(PPh₃)₂Cl₂ (3.4 g, 4.86 mmol) and Et₃N (14 mL, 6.9 mmol) in toluene(100 mL) and water (50 mL) was heated at 70° C. for 4 h. The aqueouslayer was separated and the organic layer was washed with water (80mL×2) and brine (100 mL), dried over Na₂SO₄ and concentrated underreduced pressure to dryness. The residue was recrystallized with etherto afford 2-(3,3-dimethylbut-1-ynyl)-5-fluoro-4-nitroaniline (4.2 g,46%). ¹H-NMR (400 MHz, DMSO-d₆) δ 7.84 (d, J=8.4 Hz, 1H), 6.84 (brs,2H), 6.54 (d, J=14.4 Hz, 1H), 1.29 (s, 9H).

N-(2-(3,3-Dimethylbut-1-ynyl)-5-fluoro-4-nitrophenyl)butyramide

To a solution of 2-(3,3-dimethylbut-1-ynyl)-5-fluoro-4-nitroaniline (4.2g, 17.8 mmol) in dichloromethane (50 mL) and Et₃N (10.3 mL, 71.2 mmol)was added butyryl chloride (1.9 g, 17.8 mmol) at 0° C. The mixture wasstirred at room temperature for 1 h and then poured into water. Theaqueous phase was separated and the organic layer was washed with water(50 mL×2) and brine (100 mL), dried over Na₂SO₄ and concentrated underreduced pressure to dryness. The residue was washed with ether to giveN-(2-(3,3-dimethylbut-1-ynyl)-5-fluoro-4-nitrophenyl)butyramide (3.5 g,67%), which was used in the next step without further purification.

2-tert-Butyl-6-fluoro-5-nitro-1H-indole

A solution ofN-(2-(3,3-dimethylbut-1-ynyl)-5-fluoro-4-nitrophenyl)butyramide (3.0 g,9.8 mmol) and TBAF (4.5 g, 17.2 mmol) in DMF (25 mL) was heated at 100°C. overnight. The mixture was poured into water and then extracted withEtOAc (80 mL×3). The combined extracts were washed with water (50 mL)and brine (50 mL), dried over Na₂SO₄ and concentrated under reducedpressure to dryness. The residue was purified by column chromatographyon silica gel (petroleum ether/ethyl acetate 20:1) to give compound2-tert-butyl-6-fluoro-5-nitro-1H-indole (1.5 g, 65%). H-NMR (400 MHz,CDCl₃) δ 8.30 (d, J=7.2 Hz, 1H), 7.12 (d, J=11.6 Hz, 1H), 6.35 (d, J=1.2Hz, 1H), 1.40 (s, 9H).

2-tert-Butyl-6-fluoro-1H-indol-5-amine

A suspension of 2-tert-butyl-6-fluoro-5-nitro-1H-indole (1.5 g, 6.36mmol) and Ni (0.5 g) in MeOH (20 mL) was stirred under H₂ atmosphere (1atm) at the room temperature for 3 h. The catalyst was filtered off andthe filtrate was concentrated under reduced pressure to dryness. Theresidue was recrystallized in ether to give2-tert-butyl-6-fluoro-1H-indol-5-amine (520 mg, 38%). ¹H-NMR (300 MHz,DMSO-d₆) δ 10.46 (brs, 1H), 6.90 (d, J=8.7 Hz, 1H), 6.75 (d, J=9.0 Hz,1H), 5.86 (s, 1H), 4.37 (brs, 2H), 1.29 (s, 9H); MS (ESI) m/e 206.6.

6-tert-Butoxy-2-tert-butyl-5-nitro-1H-indole

A solution ofN-(2-(3,3-dimethylbut-1-ynyl)-5-fluoro-4-nitrophenyl)butyramide (500 mg,1.63 mmol) and t-BuOK (0.37 g, 3.26 mmol) in DMF (10 mL) was heated at70° C. for 2 h. The mixture was poured into water and then extractedwith EtOAc (50 mL×3). The combined extracts were washed with water (50mL) and brine (50 mL), dried over Na₂SO₄ and concentrated under reducedpressure to give 6-tert-butoxy-2-tert-butyl-5-nitro-1H-indole (100 mg,21%). ¹H-NMR (300 MHz, DMSO-d₆) δ 11.35 (brs, 1H), 7.99 (s, 1H), 7.08(s, 1H), 6.25 (s, 1H), 1.34 (s, 9H), 1.30 (s, 9H).

6-tert-Butoxy-2-tert-butyl-1H-indol-5-amine

A suspension of 6-tert-butoxy-2-tert-butyl-5-nitro-1H-indole (100 mg,0.36 mmol) and Raney Ni (0.5 g) in MeOH (15 mL) was stirred under H₂atmosphere (1 atm) at the room temperature for 2.5 h. The catalyst wasfiltered off and the filtrate was concentrated under reduced pressure todryness. The residue was recrystallized in ether to give6-tert-butoxy-2-tert-butyl-1H-indol-5-amine (30 mg, 32%). ¹H-NMR (300MHz, MeOD) 6.98 (s, 1H), 6.90 (s, 1H), 5.94 (d, J=0.6 Hz, 1H), 1.42 (s,9H), 1.36 (s, 9H); MS (ESI) m/e 205.0.

Example 31: 1-tert-Butyl-1H-indol-5-amine

N-tert-Butyl-4-nitroaniline

A solution of 1-fluoro-4-nitro-benzene (1 g, 7.1 mmol) andtert-butylamine (1.5 g, 21 mmol) in DMSO (5 mL) was stirred at 75° C.overnight. The mixture was poured into water (10 mL) and extracted withEtOAc (7 mL×3). The combined organic layers were washed with water,brine, dried over Na₂SO₄ and concentrated under vacuum to dryness. Theresidue was purified by column chromatography on silica gel (petroleumether/ethyl acetate 30:1) to afford N-tert-butyl-4-nitroaniline (1 g,73%). ¹H NMR (CDCl₃, 400 MHz) δ 8.03-8.00 (m, 2H), 6.61-6.57 (m, 2H),4.67 (brs, 1H), 1.42 (s, 9H).

(2-Bromo-4-nitro-phenyl)-tert-butyl-amine

To a solution of N-tert-butyl-4-nitroaniline (1 g, 5.1 mmol) in AcOH (5mL) was added Br₂ (0.86 g, 54 mmol) dropwise at 15° C. After addition,the mixture was stirred at 30° C. for 30 min and then filtered. Thefilter cake was basified to pH 8-9 with aqueous NaHCO₃. The aqueouslayer was extracted with EtOAc (10 mL×3). The combined organic layerswere washed with water, brine, dried over Na₂SO₄ and concentrated undervacuum to give (2-bromo-4-nitro-phenyl)-tert-butyl-amine (0.6 g, 43%).¹H-NMR (CDCl₃, 400 MHz) δ 8.37 (dd, J=2.4 Hz, 1H), 8.07 (dd, J=2.4, 9.2Hz, 1H), 6.86 (d, J=9.2 Hz, 1H), 5.19 (brs, 1H), 1.48 (s, 9H).

tert-Butyl-(4-nitro-2-trimethylsilanylethynyl-phenyl)-amine

To a solution of (2-bromo-4-nitro-phenyl)-tert-butyl-amine (0.6 g, 2.2mmol) in Et₃N (10 mL) was added Pd(PPh₃)₂Cl₂ (70 mg, 0.1 mmol), CuI(20.9 mg, 0.1 mmol) and ethynyl-trimethyl-silane (0.32 g, 3.3 mmol)successively under N₂ protection. The reaction mixture was heated at 70°C. overnight. The solvent was removed under vacuum and the residue waswashed with EtOAc (10 mL×3). The combined organic layers were washedwith water, brine, dried over Na₂SO₄ and concentrated under vacuum todryness. The residue was purified by column chromatography on silica gel(petroleum ether/ethyl acetate 20:1) to affordtert-butyl-(4-nitro-2-trimethylsilanylethynyl-phenyl)-amine (100 mg,16%). ¹H-NMR (CDCl₃, 400 MHz) δ 8.20 (d, J=2.4, Hz, 1H), 8.04 (dd,J=2.4, 9.2 Hz, 1H), 6.79 (d, J=9.6 Hz, 1H), 5.62 (brs, 1H), 1.41 (s,9H), 0.28 (s, 9H).

1-tert-Butyl-5-nitro-1H-indole

To a solution oftert-butyl-(4-nitro-2-trimethylsilanylethynyl-phenyl)-amine (10 mg,0.035 mmol) in DMF (2 mL), was added CuI (13 mg, 0.07 mmol) under N₂protection. The reaction mixture was stirred at 100° C. overnight. Atthis time, EtOAc (4 mL) was added to the mixture. The mixture wasfiltered and the filtrate was washed with water, brine, dried overNa₂SO₄ and concentrated under vacuum to obtain1-tert-butyl-5-nitro-1H-indole (7 mg, 93%). ¹H-NMR (CDCl₃, 300 MHz) δ8.57 (d, J=2.1 Hz, 1H), 8.06 (dd, J=2.4, 9.3 Hz, 1H), 7.65 (d, J=9.3 Hz,1H), 7.43 (d, J=3.3 Hz, 1H), 6.63 (d, J=3.3 Hz, 1H), 1.76 (s, 9H).

1-tert-Butyl-1H-indol-5-amine

To a solution of 1-tert-butyl-5-nitro-1H-indole (6.5 g, 0.030 mol) inMeOH (100 mL) was added Raney Nickel (0.65 g, 10%) under N₂ protection.The mixture was stirred under hydrogen atmosphere (1 atm) at 30° C. for1 h. The catalyst was filtered off and the filtrate was concentratedunder vacuum to dryness. The residue was purified by columnchromatography on silica gel (PE/EtOAc 1:2) to give1-tert-butyl-1H-indol-5-amine (2.5 g, 45%). ¹H-NMR (CDCl₃, 400 MHz) δ7.44 (d, J=8.8 Hz, 1H), 7.19 (dd, J=3.2 Hz, 1H), 6.96 (d, J=2.0 Hz, 1H),6.66 (d, J=2.0, 8.8 Hz, 1H), 6.26 (d, J=3.2 Hz, 1H), 1.67 (s, 9H). MS(ESI) m/e (M+H⁺) 189.2.

Example 32: 2-tert-Butyl-1-methyl-1H-indol-5-amine

(2-Bromo-4-nitro-phenyl)-methyl-amine

To a solution of methyl-(4-nitro-phenyl)-amine (15.2 g, 0.1 mol) in AcOH(150 mL) and CHCl₃ (50 mL) was added Br₂ (16.0 g, 0.1 mol) dropwise at5° C. The mixture was stirred at 10° C. for 1 h and then basified withsat. aq. NaHCO₃. The resulting mixture was extracted with EtOAc (100mL×3), and the combined organics were dried over anhydrous Na₂SO₄ andevaporated under vacuum to give (2-bromo-4-nitro-phenyl)-methyl-amine(2-bromo-4-nitro-phenyl)-methyl-amine (23.0 g, 99%), which was used inthe next step without further purification. ¹H NMR (300 MHz, CDCl₃) δ8.37 (d, J=2.4 Hz, 1H), 8.13 (dd, J=2.4, 9.0 Hz, 1H), 6.58 (d, J=9.0 Hz,1H), 5.17 (brs, 1H), 3.01 (d, J=5.4 Hz, 3H).

[2-(3,3-Dimethyl-but-1-ynyl)-4-nitro-phenyl]-methyl-amine

To a solution of (2-bromo-4-nitro-phenyl)-methyl-amine (22.5 g, 97.4mmol) in toluene (200 mL) and water (100 mL) were added Et₃N (19.7 g,195 mmol), Pd(PPh₃)₂Cl₂ (6.8 g, 9.7 mmol), CuI (0.7 g, 3.9 mmol) and3,3-dimethyl-but-1-yne (16.0 g, 195 mmol) successively under N₂protection. The mixture was heated at 70° C. for 3 hours and then cooleddown to room temperature. The resulting mixture was extracted with EtOAc(100 mL×3). The combined organic extracts were dried over anhydrousNa₂SO₄ and evaporated under vacuum to give[2-(3,3-dimethyl-but-1-ynyl)-4-nitro-phenyl]-methyl-amine (20.1 g, 94%),which was used in the next step without further purification. ¹H NMR(400 MHz, CDCl₃) δ 8.15 (d, J=2.4 Hz, 1H), 8.08 (dd, J=2.8, 9.2 Hz, 1H),6.50 (d, J=9.2 Hz, 1H), 5.30 (brs, 1H), 3.00 (s, 3H), 1.35 (s, 9H).

2-tert-Butyl-1-methyl-5-nitro-1H-indole

A solution of [2-(3,3-dimethyl-but-1-ynyl)-4-nitro-phenyl]-methyl-amine(5.0 g, 22.9 mmol) and TBAF (23.9 g, 91.6 mmol) in THF (50 mL) washeated at reflux overnight. The solvent was removed by evaporation undervacuum and the residue was dissolved in brine (100 mL) and EtOAc (100mL). The organic phase was separated, dried over Na₂SO₄ and evaporatedunder vacuum to give 2-tert-butyl-1-methyl-5-nitro-1H-indole (5.0 g,99%), which was used in the next step without further purification. ¹HNMR (CDCl₃, 400 MHz) δ 8.47 (d, J=2.4 Hz, 1H), 8.07 (dd, J=2.4, 9.2 Hz,1H), 7.26-7.28 (m, 1H), 6.47 (s, 1H), 3.94 (s, 3H), 1.50 (s, 9H).

2-tert-Butyl-1-methyl-1H-indol-5-amine

To a solution of 2-tert-butyl-1-methyl-5-nitro-1H-indole (3.00 g, 13.7mmol) in MeOH (30 mL) was added Raney Ni (0.3 g) under nitrogenatmosphere. The mixture was stirred under hydrogen atmosphere (1 atm) atroom temperature overnight. The mixture was filtered through a Celitepad and the filtrate was evaporated under vacuum. The crude residue waspurified by column chromatography on silica gel (P.E/EtOAc 20:1) to give2-tert-butyl-1-methyl-1H-indol-5-amine (1.7 g, 66%). ¹H NMR (300 MHz,CDCl₃) δ 7.09 (d, J=8.4 Hz, 1H), 6.89-6.9 (m, 1H), 6.66 (dd, J=2.4, 8.7Hz, 1H), 6.14 (d, J=0.6 Hz, 1H), 3.83 (s, 3H), 3.40 (brs, 2H), 1.45 (s,9H); MS (ESI) m/e (M+H⁺) 203.1.

Example 33: 2-Cyclopropyl-1H-indol-5-amine

2-Bromo-4-nitroaniline

To a solution of 4-nitro-aniline (25 g, 0.18 mol) in HOAc (150 mL) wasadded liquid Br₂ (30 g, 0.19 mol) dropwise at room temperature. Themixture was stirred for 2 hours. The solid was collected by filtrationand poured into water (100 mL), which was basified with sat. aq. NaHCO₃to pH 7 and extracted with EtOAc (300 mL×3). The combined organic layerswere dried over anhydrous Na₂SO₄ and evaporated under reduced pressureto give 2-bromo-4-nitroaniline (30 g, 80%), which was directly used inthe next step.

2-(Cyclopropylethynyl)-4-nitroaniline

To a deoxygenated solution of 2-bromo-4-nitroaniline (2.17 g, 0.01mmol), ethynyl-cyclopropane (1 g, 15 mmol) and CuI (10 mg, 0.05 mmol) intriethylamine (20 mL) was added Pd(PPh₃)₂Cl₂ (210 mg, 0.3 mmol) underN₂. The mixture was heated at 70° C. and stirred for 24 hours. The solidwas filtered off and washed with EtOAc (50 mL×3). The filtrate wasevaporated under reduced pressure, and the residue was purified bycolumn chromatography on silica gel (petroleum ether/ethyl acetate=10/1)to give 2-(cyclopropylethynyl)-4-nitroaniline (470 mg, 23%). ¹H NMR (300MHz, CDCl₃) δ 8.14 (d, J=2.7 Hz, 1H), 7.97 (dd, J=2.7, 9.0 Hz, 1H), 6.63(d, J=9.0 Hz, 1H), 4.81 (brs, 2H), 1.55-1.46 (m, 1H), 0.98-0.90 (m, 2H),0.89-0.84 (m, 2H).

N-(2-(Cyclopropylethynyl)phenyl)-4-nitrobutyramide

To a solution of 2-(cyclopropylethynyl)-4-nitroaniline (3.2 g, 15.8mmol) and pyridine (2.47 g, 31.7 mmol) in CH₂Cl₂ (60 mL) was addedbutyryl chloride (2.54 g, 23.8 mmol) at 0° C. The mixture was warmed toroom temperature and stirred for 3 hours. The resulting mixture waspoured into ice-water. The organic layer was separated. The aqueousphase was extracted with CH₂Cl₂ (30 m L×3). The combined organic layerswere dried over anhydrous Na₂SO₄ and evaporated under reduced pressureto give the crude product, which was purified by column chromatographyon silica gel (petroleum ether/ethyl acetate=10/1) to giveN-(2-(cyclopropylethynyl)phenyl)-4-nitrobutyramide (3.3 g, 76%). ¹H NMR(400 MHz, CDCl₃) δ 8.61 (d, J=9.2 Hz, 1H), 8.22 (d, J=2.8 Hz, 1H), 8.18(brs, 1H), 8.13 (dd, J=2.4, 9.2 Hz, 1H), 2.46 (t, J=7.2 Hz, 2H),1.83-1.76 (m, 2H), 1.59-1.53 (m, 1H), 1.06 (t, J=7.2 Hz, 3H), 1.03-1.01(m, 2H), 0.91-0.87 (m, 2H).

2-Cyclopropyl-5-nitro-1H-indole

A mixture of N-(2-(cyclopropylethynyl)phenyl)-4-nitrobutyramide (3.3 g,0.01 mol) and TBAF (9.5 g, 0.04 mol) in THF (100 mL) was heated atreflux for 24 hours. The mixture was cooled to the room temperature andpoured into ice water. The mixture was extracted with CH₂Cl₂ (50 m L×3).The combined organic layers were dried over anhydrous Na₂SO₄ andevaporated under reduced pressure. The residue was purified by columnchromatography on silica gel (petroleum ether/ethyl acetate=10/1) togive 2-cyclopropyl-5-nitro-1H-indole (1.3 g, 64%). ¹H NMR (400 MHz,CDCl₃) δ 8.44 (d, J=2.0 Hz, 1H), 8.40 (brs, 1H), 8.03 (dd, J=2.0, 8.8Hz, 1H), 7.30 (d, J=8.8 Hz, 1H), 6.29 (d, J=0.8 Hz, 1H), 2.02-1.96 (m,1H) 1.07-1.02 (m, 2H), 0.85-0.81 (m, 2H).

2-Cyclopropyl-1H-indol-5-amine

To a solution of 2-cyclopropyl-5-nitro-1H-indole (1.3 g, 6.4 mmol) inMeOH (30 mL) was added Raney Nickel (0.3 g) under nitrogen atmosphere.The mixture was stirred under hydrogen atmosphere (1 atm) at roomtemperature overnight. The catalyst was filtered through a Celite padand the filtrate was evaporated under vacuum to give the crude product,which was purified by column chromatography on silica gel (petroleumether/ethyl acetate=5/1) to give 2-cyclopropyl-1H-indol-5-amine (510 mg,56%). ¹H NMR (400 MHz, CDCl₃) δ 6.89 (d, J=8.4 Hz, 1H), 6.50 (d, J=1.6Hz, 1H), 6.33 (dd, J=2.0, 8.4 Hz, 1H), 5.76 (s, 1H), 4.33 (brs, 2H),1.91-1.87 (m, 1H), 0.90-0.85 (m, 2H), 0.70-0.66 (m, 2H); MS (ESI) m/e(M+H⁺) 173.2.

Example 34: 3-tert-Butyl-1H-indol-5-amine

3-tert-Butyl-5-nitro-1H-indole

To a mixture of 5-nitro-1H-indole (6 g, 36.8 mmol) and AlCl₃ (24 g, 0.18mol) in CH₂Cl₂ (100 mL) was added 2-bromo-2-methyl-propane (8.1 g, 36.8mmol) dropwise at 0° C. After being stirred at 15° C. overnight, thereaction mixture was poured into ice (100 mL). The precipitated saltswere removed by filtration and the aqueous layer was extracted withCH₂Cl₂ (30 mL×3). The combined organic layers were washed with water,brine, dried over Na₂SO₄ and concentrated under vacuum to obtain thecrude product, which was purified by column chromatography on silica gel(petroleum ether/ethyl acetate 20:1) to give3-tert-butyl-5-nitro-1H-indole (2.5 g, 31%). ¹H NMR (CDCl₃ 400 MHz) δ8.49 (d, J=1.6 Hz, 1H), 8.31 (brs, 1H), 8.05 (dd, J=2.0, 8.8 Hz, 1H),7.33 (d, J=8.8 Hz, 1H), 6.42 (d, J=1.6 Hz, 1H), 1.42 (s, 9H).

3-tert-Butyl-1H-indol-5-amine

To a solution of 3-tert-butyl-5-nitro-1H-indole (2.5 g, 11.6 mmol) inMeOH (30 mL) was added Raney Nickel (0.2 g) under N₂ protection. Themixture was stirred under hydrogen atmosphere (1 atm) at 15° C. for 1hr. The catalyst was filtered off and the filtrate was concentratedunder vacuum to dryness. The residue was purified by preparative HLPC toafford 3-tert-butyl-1H-indol-5-amine (0.43 g, 19%). ¹H NMR (CDCl₃, 400MHz) δ 7.72 (brs, 1H), 7.11 (d, J=8.4 Hz, 1H), 6.86 (d, J=2.0 Hz, 1H),6.59 (dd, J=2.0, 8.4 Hz, 1H), 6.09 (d, J=1.6 Hz, 1H), 1.37 (s, 9H); MS(ESI) m/e (M+H⁺) 189.1.

Example 35: 2-Phenyl-1H-indol-5-amine

2-Bromo-4-nitroaniline

To a solution of 4-nitroaniline (50 g, 0.36 mol) in AcOH (500 mL) wasadded liquid Br₂ (60 g, 0.38 mol) dropwise at 5° C. The mixture wasstirred for 30 min at that temperature. The insoluble solid wascollected by filtration and poured into EtOAc (200 mL). The mixture wasbasified with saturated aqueous NaHCO₃ to pH 7. The organic layer wasseparated. The aqueous phase was extracted with EtOAc (300 mL×3). Thecombined organic layers were dried and evaporated under reduced pressureto give 2-bromo-4-nitroaniline (56 g, 72%), which was directly used inthe next step.

4-Nitro-2-(phenylethynyl)aniline

To a deoxygenated solution of 2-bromo-4-nitroaniline (2.17 g, 0.01mmol), ethynyl-benzene (1.53 g, 0.015 mol) and CuI (10 mg, 0.05 mmol) intriethylamine (20 mL) was added Pd(PPh₃)₂Cl₂ (210 mg, 0.2 mmol) underN₂. The mixture was heated at 70° C. and stirred for 24 hours. The solidwas filtered off and washed with EtOAc (50 mL×3). The filtrate wasevaporated under reduced pressure and the residue was purified by columnchromatography on silica gel (petroleum ether/ethyl acetate=10/1) togive 4-nitro-2-(phenylethynyl)aniline (340 mg, 14%). ¹H NMR (300 MHz,CDCl₃) δ 8.37-8.29 (m, 1H), 8.08-8.00 (m, 1H), 7.56-7.51 (m, 2H),7.41-7.37 (m, 3H), 6.72 (m, 1H), 4.95 (brs, 2H).

N-(2-(Phenylethynyl)phenyl)-4-nitrobutyramide

To a solution of 4-nitro-2-(phenylethynyl)aniline (17 g, 0.07 mmol) andpyridine (11.1 g, 0.14 mol) in CH₂Cl₂ (100 mL) was added butyrylchloride (11.5 g, 0.1 mol) at 0° C. The mixture was warmed to roomtemperature and stirred for 3 hours. The resulting mixture was pouredinto ice-water. The organic layer was separated. The aqueous phase wasextracted with CH₂Cl₂ (30 m L×3). The combined organic layers were driedover anhydrous Na₂SO₄ and evaporated under reduced pressure. The residuewas purified by column chromatography on silica gel (petroleumether/ethyl acetate=10/1) to giveN-(2-(phenylethynyl)phenyl)-4-nitrobutyramide (12 g, 55%). ¹H NMR (400MHz, CDCl₃) δ 8.69 (d, J=9.2 Hz, 1H), 8.39 (d, J=2.8 Hz, 1H), 8.25-8.20(m, 2H), 7.58-7.55 (m, 2H), 7.45-7.42 (m, 3H), 2.49 (t, J=7.2 Hz, 2H),1.85-1.79 (m, 2H), 1.06 (t, J=7.2 Hz, 3H).

5-Nitro-2-phenyl-1H-indole

A mixture of N-(2-(phenylethynyl)phenyl)-4-nitrobutyramide (5.0 g, 0.020mol) and TBAF (12.7 g, 0.050 mol) in THF (30 mL) was heated at refluxfor 24 h. The mixture was cooled to room temperature and poured into icewater. The mixture was extracted with CH₂Cl₂ (50 m L×3). The combinedorganic layers were dried over anhydrous Na₂SO₄ and evaporated underreduced pressure. The residue was purified by column chromatography onsilica gel (petroleum ether/ethyl acetate=10/1) to give5-nitro-2-phenyl-1H-indole (3.3 g, 69%). ¹H NMR (400 MHz, CDCl₃) δ 8.67(s, 1H), 8.06 (dd, J=2.0, 8.8 Hz, 1H), 7.75 (d, J=7.6 Hz, 2H), 7.54 (d,J=8.8 Hz, 1H), 7.45 (t, J=7.6 Hz, 2H), 7.36 (t, J=7.6 Hz, 1H). 6.95 (s,1H).

2-Phenyl-1H-indol-5-amine

To a solution of 5-nitro-2-phenyl-1H-indole (2.83 g, 0.01 mol) in MeOH(30 mL) was added Raney Ni (510 mg) under nitrogen atmosphere. Themixture was stirred under hydrogen atmosphere (1 atm) at roomtemperature overnight. The catalyst was filtered through a Celite padand the filtrate was evaporated under vacuum to give the crude product,which was purified by column chromatography on silica gel (petroleumether/ethyl acetate=5/1) to give 2-phenyl-1H-indol-5-amine (1.6 g, 77%).¹H NMR (400 MHz, CDCl₃) δ 7.76 (d, J=7.6 Hz, 2H), 7.39 (t, J=7.6 Hz,2H), 7.24 (t, J=7.6 Hz, 1H), 7.07 (d, J=8.4 Hz, 1H), 6.64 (d, J=1.6 Hz,1H), 6.60 (d, J=1.2 Hz, 1H), 6.48 (dd, J=2.0, 8.4 Hz, 1H), 4.48 (brs,2H); MS (ESI) m/e (M+H⁺) 209.0.

Example 36: 2-tert-Butyl-4-fluoro-1H-indol-5-amine

2-Bromo-3-fluoroaniline

To a solution of 2-bromo-1-fluoro-3-nitrobenzene (1.0 g, 5.0 mmol) inCH₃OH (50 mL) was added NiCl₂ (2.2 g 10 mmol) and NaBH₄ (0.50 g 14 mmol)at 0° C. After the addition, the mixture was stirred for 5 min. Water(20 mL) was added and the mixture was extracted with EtOAc (20 mL×3).The organic layers were dried over anhydrous Na₂SO₄ and evaporated undervacuum to give 2-bromo-3-fluoroaniline (600 mg, 70%). ¹H NMR (400 MHz,CDCl₃) δ 7.07-7.02 (m, 1H), 6.55-6.49 (m, 1H), 4.22 (br s, 2H).

N-(2-Bromo-3-fluorophenyl)butyramide

To a solution of 2-bromo-3-fluoroaniline (2.0 g, 11 mmol) in CH₂Cl₂ (50mL) was added butyryl chloride (1.3 g, 13 mmol) and pyridine (1.7 g, 21mmol) at 0° C. The mixture was stirred at room temperature for 24 h.Water (20 mL) was added and the mixture was extracted with CH₂Cl₂ (50mL×3). The organic layers were dried anhydrous over Na₂SO₄ andevaporated under vacuum to give N-(2-bromo-3-fluorophenyl)butyramide(2.0 g, 73%), which was directly used in the next step.

N-(2-(3,3-Dimethylbut-1-ynyl)-3-fluorophenyl)butyramide

To a solution of N-(2-bromo-3-fluorophenyl)butyramide (2.0 g, 7.0 mmol)in Et₃N (100 mL) was added 4,4-dimethylpent-2-yne (6.0 g, 60 mmol), CuI(70 mg, 3.8 mmol), and Pd(PPh₃)₂Cl₂ (500 mg) successively at roomtemperature under N₂. The mixture was heated at 80° C. overnight. Thecooled mixture was filtered and the filtrate was extracted with EtOAc(40 mL×3). The organic layers were washed with sat. NaCl, dried overanhydrous Na₂SO₄, and evaporated under vacuum. The crude compound waspurified by column chromatography on silica gel (10% EtOAc in petroleumether) to give N-(2-(3,3-dimethylbut-1-ynyl)-3-fluorophenyl)butyramide(1.1 g, 55%). ¹H NMR (400 MHz, CDCl₃) δ 8.20 (d, J=7.6, 1H), 7.95 (s,1H), 7.21 (m, 1H), 6.77 (t, J=7.6 Hz, 1H), 2.39 (t, J=7.6 Hz, 2H),1.82-1.75 (m, 2H), 1.40 (s, 9H), 1.12 (t, J=7.2 Hz, 3H).

2-tert-Butyl-4-fluoro-1H-indole

To a solution of N-(2-(3,3-dimethylbut-1-ynyl)-3-fluorophenyl)butyramide(6.0 g, 20 mmol) in DMF (100 mL) was added t-BuOK (5.0 g, 50 mmol) atroom temperature. The mixture was heated at 90° C. overnight before itwas poured into water and extracted with EtOAc (100 mL×3). The organiclayers were washed with sat. NaCl and water, dried over anhydrousNa₂SO₄, and evaporated under vacuum to give2-tert-butyl-4-fluoro-1H-indole (5.8 g, 97%). ¹H NMR (400 MHz, CDCl₃) δ8.17 (br s, 1H), 7.11 (d, J=7.2 Hz, 1H), 7.05-6.99 (m, 1H), 6.76-6.71(m, 1H), 6.34 (m, 1H), 1.41 (s, 9H).

2-tert-Butyl-4-fluoro-5-nitro-1H-indole

To a solution of 2-tert-butyl-4-fluoro-1H-indole (2.5 g, 10 mmol) inH₂SO₄ (30 mL) was added KNO₃ (1.3 g, 10 mmol) at 0° C. The mixture wasstirred for 0.5 h at −10° C. The mixture was poured into water andextracted with EtOAc (100 mL×3). The organic layers were washed withsat. NaCl and water, dried over anhydrous Na₂SO₄, and evaporated undervacuum. The crude compound was purified by column chromatography onsilica gel (10% EtOAc in petroleum ether) to give2-tert-butyl-4-fluoro-5-nitro-1H-indole (900 mg, 73%). ¹H NMR (400 MHz,CDCl₃) δ 8.50 (br s, 1H), 7.86 (dd, J=7.6, 8.8 Hz, 1H), 7.13 (d, J=8.8Hz, 1H), 6.52 (dd, J=0.4, 2.0 Hz, 1H), 1.40 (s, 9H).

2-tert-Butyl-4-fluoro-1H-indol-5-amine

To a solution of 2-tert-butyl-4-fluoro-5-nitro-1H-indole (2.1 g, 9.0mmol) in methanol (50 mL) was added NiCl₂ (4.2 g, 18 mmol) and NaBH₄(1.0 g, 27 mmol) at 0° C. After the addition, the mixture was stirredfor 5 min. Water (20 mL) was added and the mixture was extracted withEtOAc (30 mL×3). The organic layers were washed with sat. NaCl andwater, dried over anhydrous Na₂SO₄, evaporated under vacuum to give2-tert-butyl-4-fluoro-1H-indol-5-amine (900 mg, 50%). ¹H NMR (300 MHz,CDCl₃) δ 7.80 (brs, 1H), 6.91 (d, J=8.4 Hz, 1H), 6.64 (dd, J=0.9, 2.4Hz, 1H), 6.23 (s, 1H), 1.38 (s, 9H).

Example 37: 2,3,4,9-Tetrahydro-1H-carbazol-6-amine

2,3,4,9-Tetrahydro-1H-carbazol-6-amine

6-Nitro-2,3,4,9-tetrahydro-1H-carbazole (0.100 g, 0.462 mmol) wasdissolved in a 40 mL scintillation vial containing a magnetic stir barand 2 mL of ethanol. Tin(II) chloride dihydrate (1.04 g, 4.62 mmol) wasadded to the reaction mixture and the resulting suspension was heated at70° C. for 16 h. The crude reaction mixture was then diluted with 15 mLof a saturated aqueous solution of sodium bicarbonate and extractedthree times with an equivalent volume of ethyl acetate. The ethylacetate extracts were combined, dried over sodium sulfate, andevaporated to dryness to yield 2,3,4,9-tetrahydro-1H-carbazol-6-amine(82 mg, 95%) which was used without further purification.

Example 38: 2-tert-Butyl-7-fluoro-1H-indol-5-amine

2-Bromo-6-fluoro-4-nitro-phenylamine

To a solution of 2-fluoro-4-nitro-phenylamine (12 g, 77 mmol) in AcOH(50 mL) was added Br₂ (3.9 mL, 77 mmol) dropwise at 0° C. The mixturewas stirred at 20° C. for 3 h. The reaction mixture was basified withsat. aq. NaHCO₃, and extracted with EtOAc (100 mL×3). The combinedorganics were dried over anhydrous Na₂SO₄ and evaporated under vacuum togive 2-bromo-6-fluoro-4-nitro-phenylamine (18 g, 97%). ¹H NMR (400 MHz,CDCl₃) δ 8.22 (m, 1H), 7.90 (dd, J=2.4, 10.8 Hz, 1H), 4.88 (brs, 2H).

2-(3,3-Dimethyl-but-1-ynyl)-6-fluoro-4-nitro-phenylamine

To a solution of 2-bromo-6-fluoro-4-nitro-phenylamine (11 g, 47 mmol) indry Et₃N (100 mL) was added CuI (445 mg, 5% mol), Pd(PPh₃)₂Cl₂ (550 mg,5% mol) and 3,3-dimethyl-but-1-yne (9.6 g, 120 mmol) under N₂protection. The mixture was stirred at 80° C. for 10 h. The reactionmixture was filtered, poured into ice (100 g), and extracted with EtOAc(50 mL×3). The combined organic extracts were dried over anhydrousNa₂SO₄ and evaporated under vacuum to give the crude product, which waspurified by column chromatography on silica gel (petroleum ether/ethylacetate 50:1) to give2-(3,3-dimethyl-but-1-ynyl)-6-fluoro-4-nitro-phenylamine (4.0 g, 36%).¹H NMR (400 MHz, CDCl₃) δ 8.02 (d, J=1.2 Hz, 1H), 7.84 (dd, J=2.4, 10.8Hz, 1H), 4.85 (brs, 2H), 1.36 (s, 9H).

N-[2-(3,3-Dimethyl-but-1-ynyl)-6-fluoro-4-nitro-phenyl]-butyramide

To a solution of2-(3,3-dimethyl-but-1-ynyl)-6-fluoro-4-nitro-phenylamine (4.0 g, 17mmol) and pyridine (2.7 g, 34 mmol) in anhydrous CH₂Cl₂ (30 mL) wasadded and butyryl chloride (1.8 g, 17 mmol) dropwise at 0° C. Afterstirring for 5 h at 0° C., the reaction mixture was poured into ice (50g) and extracted with CH₂Cl₂ (30 mL×3). The combined organic extractswere dried over anhydrous Na₂SO₄ and evaporated under vacuum to giveN-[2-(3,3-dimethyl-but-1-ynyl)-6-fluoro-4-nitro-phenyl]-butyramide (3.2g, 62%), which was used in the next step without further purification.¹H NMR (300 MHz, DMSO) δ 8.10 (dd, J=1.5, 2.7 Hz, 1H), 7.95 (dd, J=2.4,9.6 Hz, 1H), 7.22 (brs, 1H), 2.45 (t, J=7.5 Hz, 2H), 1.82 (m, 2H), 1.36(s, 9H), 1.06 (t, J=7.5 Hz, 3H).

2-tert-Butyl-7-fluoro-5-nitro-1H-indole

To a solution ofN-[2-(3,3-dimethyl-but-1-ynyl)-6-fluoro-4-nitro-phenyl]-butyramide (3.2g, 10 mmol) in DMF (20 mL) was added t-BuOK (2.3 g, 21 mmol) at roomtemperature. The mixture was heated at 120° C. for 2 g before beingcooled down to room temperature. Water (50 mL) was added to the reactionmixture and the resulting mixture was extracted with CH₂Cl₂ (30 mL×3).The combined organic extracts were dried over anhydrous Na₂SO₄ andevaporated under vacuum to give 2-tert-butyl-7-fluoro-5-nitro-1H-indole(2.0 g, 81%), which was used in the next step without furtherpurification. ¹H NMR (300 MHz, CDCl₃) δ 9.95 (brs, 1H), 8.30 (d, J=2.1Hz, 1H), 7.74 (dd, J=1.8, 11.1 Hz, 1H), 6.43 (dd, J=2.4, 3.3 Hz, 1H),1.43 (s, 9H).

2-tert-Butyl-7-fluoro-1H-indol-5-amine

To a solution of 2-tert-butyl-7-fluoro-5-nitro-1H-indole (2.0 g, 8.5mmol) in MeOH (20 mL) was added Ni (0.3 g) under nitrogen atmosphere.The reaction mixture was stirred under hydrogen atmosphere (1 atm) atroom temperature overnight. The catalyst was filtered off through thecelite pad and the filtrate was evaporated under vacuum. The crudeproduct was purified by column chromatography on silica gel (petroleumether/ethyl acetate 100:1) to give2-tert-butyl-7-fluoro-1H-indol-5-amine (550 mg, 24%). ¹H NMR (300 MHz,CDCl₃) δ 7.87 (brs, 1H), 6.64 (d, J=1.5 Hz, 1H), 6.37 (dd, J=1.8, 12.3Hz, 1H), 6.11 (dd, J=2.4, 3.6 Hz, 1H), 1.39 (s, 9H). MS (ESI) m/z (M+H⁺)207.

Example 39: 5-Amino-2-tert-butyl-1H-indole-7-carbonitrile

2-Amino-3-(3,3-dimethylbut-1-ynyl)-5-nitrobenzonitrile

To a stirred solution of 2-amino-3-bromo-5-nitrobenzonitrile (2.4 g, 10mmol) in dry Et₃N (60 mL) was added CuI (380 mg, 5% mol) andPd(PPh₃)₂Cl₂ (470 mg, 5% mol) at room temperature.3,3-dimethyl-but-1-yne (2.1 g, 25 mmol) was added dropwise to themixture at room temperature. The reaction mixture was stirred at 80° C.for 10 h. The reaction mixture was filtered and the filtrate was pouredinto ice (60 g), extracted with ethyl acetate. The phases were separatedand the organic phase was dried over Na₂SO₄. The solvent was removedunder vacuum to obtain the crude product, which was purified by columnchromatography (2-10% EtOAc in petroleum ether) to obtain2-amino-3-(3,3-dimethylbut-1-ynyl)-5-nitrobenzonitrile (1.7 g, 71%). ¹HNMR (300 MHz, CDCl₃) δ 8.28 (d, J=2.7 Hz, 1H), 8.27 (d, J=2.7 Hz, 1H),5.56 (br s, 2H), 1.37 (s, 9H).

2-tert-Butyl-5-nitro-1H-indole-7-carbonitrile

To a solution of 2-amino-3-(3,3-dimethylbut-1-ynyl)-5-nitrobenzonitrile(1.7 g, 7.0 mmol) in THF (35 mL) was added TBAF (9.5 g, 28 mmol) at roomtemperature. The mixture was heated at reflux overnight. The reactionmixture was cooled and the THF was removed under reduced pressure. Water(50 ml) was added to the residue and the mixture was extracted withEtOAc. The organics were dried over Na₂SO₄ and the solvent wasevaporated under vacuum to obtain 0.87 g of crude product2-tert-butyl-5-nitro-1H-indole-7-carbonitrile which was used directly inthe next step without purification.

5-Amino-2-tert-butyl-1H-indol-7-carbonitrile

To a solution of crude product2-tert-butyl-5-nitro-1H-indole-7-carbonitrile (0.87 g, 3.6 mmol) in MeOH(10 mL) was added NiCl₂.6H₂O (1.8 g, 7.2 mmol) at −5° C. The reactionmixture was stirred for 30 min, then NaBH₄ (0.48 g, 14.32 mmol) wasadded to the reaction mixture at 0° C. After 5 min, the reaction mixturewas quenched with water, filtered and extracted with EtOAc. The combinedorganic layers were dried over Na₂SO₄ and concentrated under vacuum toobtain the crude product, which was purified by column chromatography(5-20% EtOAc in petroleum ether) to obtain5-amino-2-tert-butyl-1H-indol-7-carbonitrile (470 mg, 32% over twosteps). ¹H NMR (400 MHz, CDCl₃) δ 8.25 (s, 1H), 7.06 (d, J=2.4 Hz, 1H),6.84 (d, J=2.4 Hz, 1H), 6.14 (d, J=2.4 Hz, 1H), 3.57 (br s, 2H), 1.38(s, 9H). MS (ESI) m/z: 214 (M+H⁺).

Example 40: Methyl 5-amino-2-tert-butyl-1H-indole-7-carboxylate

2-tert-Butyl-5-nitro-1H-indole-7-carboxylic acid

2-tert-Butyl-5-nitro-1H-indole-7-carbonitrile (4.6 g, 19 mmol) was addedto a solution of KOH in EtOH (10%, 100 mL) and the mixture was heated atreflux overnight. The solution was evaporated to remove alcohol, a smallamount of water was added, and then the mixture was acidified withdilute hydrochloric acid. Upon standing in the refrigerator, anorange-yellow solid precipitated, which was purified by chromatographyon silica gel (15% EtOAc in petroleum ether) to afford2-tert-butyl-5-nitro-1H-indole-7-carboxylic acid (4.0 g, 77%). ¹H NMR(CDCl₃, 300 MHz) δ 10.79 (brs, 1H), 8.66 (s, 1H), 8.45 (s, 1H), 6.57 (s,1H), 1.39 (s, 9H).

Methyl 2-tert-butyl-5-nitro-1H-indole-7-carboxylate

SOCl₂ (3.6 g, 30 mol) was added dropwise to a solution of2-tert-butyl-5-nitro-1H-indole-7-carboxylic acid (4.0 g, 15 mol) andmethanol (30 mL) at 0° C. The mixture was stirred at 80° C. for 12 h.The solvent was evaporated under vacuum and the residue was purified bycolumn chromatography on silica gel (5% EtOAc in petroleum ether) toafford methyl 2-tert-butyl-5-nitro-1H-indole-7-carboxylate (2.95 g,70%). ¹H NMR (CDCl₃, 300 MHz) δ 9.99 (brs, 1H), 8.70 (d, J=2.1 Hz, 1H),8.65 (d, J=2.1 Hz, 1H), 6.50 (d, J=2.4 Hz, 1H), 4.04 (s, 3H), 1.44 (s,9H).

Methyl 5-amino-2-tert-butyl-1H-indole-7-carboxylate

A solution of 2-tert-butyl-5-nitro-1H-indole-7-carboxylate (2.0 g, 7.2mmol) and Raney Nickel (200 mg) in CH₃OH (50 mL) was stirred for 5 h atthe room temperature under H₂ atmosphere. The catalyst was filtered offthrough a celite pad and the filtrate was evaporated under vacuum togive methyl 5-amino-2-tert-butyl-1H-indole-7-carboxylate (1.2 g, 68%) ¹HNMR (CDCl₃, 400 MHz) δ 9.34 (brs, 1H), 7.24 (d, J=1.6 Hz, 1H), 7.10 (s,1H), 6.12 (d, J=1.6 Hz, 1H), 3.88 (s, 3H), 1.45 (s, 9H).

Example 41: (5-Amino-2-tert-butyl-1H-indol-7-yl)methanol

(2-tert-Butyl-5-nitro-1H-indol-7-yl) methanol

To a solution of methyl 2-tert-butyl-5-nitro-1H-indole-7-carboxylate(6.15 g, 22.3 mmol) and dichloromethane (30 ml) was added DIBAL-H (1.0M, 20 mL, 20 mmol) at 78° C. The mixture was stirred for 1 h beforewater (10 mL) was added slowly. The resulting mixture was extracted withEtOAc (120 mL×3). The combined organic extracts were dried overanhydrous Na₂SO₄ and evaporated under vacuum to give(2-tert-butyl-5-nitro-1H-indol-7-yl)methanol (4.0 g, 73%), which wasused in the next step directly.

(5-Amino-2-tert-butyl-1H-indol-7-yl)methanol

A mixture of (2-tert-butyl-5-nitro-1H-indol-7-yl)methanol (4.0 g, 16mmol) and Raney Nickel (400 mg) in CH₃OH (100 mL) was stirred for 5 g atroom temperature under H₂. The catalyst was filtered off through acelite pad and the filtrate was evaporated under vacuum to give(5-amino-2-tert-butyl-1H-indol-7-yl)methanol (3.4 g, 80%). ¹H NMR(CDCl₃, 400 MHz) δ 8.53 (br s, 1H), 6.80 (d, J=2.0 Hz, 1H), 6.38 (d,J=1.6 Hz, 1H), 4.89 (s, 2H), 1.37 (s, 9H).

Example 42: 2-(1-Methylcyclopropyl)-1H-indol-5-amine

Trimethyl-(1-methyl-cyclopropylethynyl)-silane

To a solution of cyclopropylethynyl-trimethyl-silane (3.0 g, 22 mmol) inether (20 mL) was added dropwise n-BuLi (8.6 mL, 21.7 mol, 2.5 Msolution in hexane) at 0° C. The reaction mixture was stirred at ambienttemperature for 24 h before dimethyl sulfate (6.85 g, 54.3 mmol) wasadded dropwise at −10° C. The resulting solution was stirred at 10° C.and then at 20° C. for 30 min each. The reaction was quenched by addinga mixture of sat. aq. NH₄Cl and 25% aq. ammonia (1:3, 100 mL). Themixture was then stirred at ambient temperature for 1 h. The aqueousphase was extracted with diethyl ether (3×50 mL) and the combinedorganic layers were washed successively with 5% aqueous hydrochloricacid (100 mL), 5% aq. NaHCO₃ solution (100 mL), and water (100 mL). Theorganics were dried over anhydrous NaSO₄ and concentrated at ambientpressure. After fractional distillation under reduced pressure,trimethyl-(1-methyl-cyclopropylethynyl)-silane (1.7 g, 52%) was obtainedas a colorless liquid. ¹H NMR (400 MHz, CDCl₃) δ 1.25 (s, 3H), 0.92-0.86(m, 2H), 0.58-0.56 (m, 2H), 0.15 (s, 9H).

1-Ethynyl-1-methyl-cyclopropane

To a solution of trimethyl-(1-methyl-cyclopropylethynyl)-silane (20 g,0.13 mol) in THF (250 mL) was added TBAF (69 g, 0.26 mol). The mixturewas stirred overnight at 20° C. The mixture was poured into water andthe organic layer was separated. The aqueous phase was extracted withTHF (50 mL). The combined organic layers were dried over anhydrousNa₂SO₄ and distilled under atmospheric pressure to obtain1-ethynyl-1-methyl-cyclopropane (7.0 g, contained 1/2 THF, 34%). ¹H NMR(400 MHz, CDCl₃) δ 1.82 (s, 1H), 1.26 (s, 3H), 0.90-0.88 (m, 2H),0.57-0.55 (m, 2H).

2-Bromo-4-nitroaniline

To a solution of 4-nitro-phenylamine (50 g, 0.36 mol) in AcOH (500 mL)was added Br₂ (60 g, 0.38 mol) dropwise at 5° C. The mixture was stirredfor 30 min at that temperature. The insoluble solid was collected byfiltration and basified with saturated aqueous NaHCO₃ to pH 7. Theaqueous phase was extracted with EtOAc (300 mL×3). The combined organiclayers were dried and evaporated under reduced pressure to obtaincompound 2-bromo-4-nitroaniline (56 g, 72%), which was directly used inthe next step.

2-((1-Methylcyclopropyl)ethynyl)-4-nitroaniline

To a deoxygenated solution of 2-bromo-4-nitroaniline (430 mg, 2.0 mmol)and 1-ethynyl-1-methyl-cyclopropane (630 mg, 8.0 mmol) in triethylamine(20 mL) was added CuI (76 mg, 0.40 mmol) and Pd(PPh₃)₂Cl₂ (140 mg, 0.20mmol) under N₂. The mixture was heated at 70° C. and stirred for 24 h.The solid was filtered off and washed with EtOAc (50 mL×3). The filtratewas evaporated under reduced pressure and the residue was purified bycolumn chromatography on silica gel (petroleum ether/ethyl acetate=10/1)to give 2-((1-methylcyclopropyl)ethynyl)-4-nitroaniline (340 mg, 79%).¹H NMR (300 MHz, CDCl₃) δ 8.15-8.14 (m, 1H), 7.98-7.95 (m, 1H), 6.63 (d,J=6.9 Hz, 1H), 4.80 (brs, 2H), 1.38 (s, 3H), 1.04-1.01 (m, 2H),0.76-0.73 (m, 2H).

N-[2-(1-Methyl-cyclopropylethynyl)-4-nitro-phenyl]-butyramide

To a solution of 2-((1-methylcyclopropyl)ethynyl)-4-nitroaniline (220mg, 1.0 mmol) and pyridine (160 mg, 2.0 mol) in CH₂Cl₂ (20 mL) was addedbutyryl chloride (140 mg, 1.3 mmol) at 0° C. The mixture was warmed toroom temperature and stirred for 3 h. The mixture was poured intoice-water. The organic layer was separated and the aqueous phase wasextracted with CH₂Cl₂ (30 mL×3). The combined organic layers were driedover anhydrous Na₂SO₄ and evaporated under reduced pressure to obtainN-[2-(1-methyl-cyclopropyl-ethynyl)-4-nitro-phenyl]-butyramide (230 mg,82%), which was directly used in the next step.

2-(1-Methylcyclopropyl)-5-nitro-1H-indole

A mixture ofN-[2-(1-methyl-cyclopropylethynyl)-4-nitro-phenyl]-butyramide (1.3 g,4.6 mmol) and TBAF (2.4 g, 9.2 mmol) in THF (20 mL) was heated at refluxfor 24 h. The mixture was cooled to room temperature and poured into icewater. The mixture was extracted with CH₂Cl₂ (30 mL×3). The combinedorganic layers were dried over anhydrous Na₂SO₄ and evaporated underreduced pressure. The residue was purified by column chromatography onsilica gel (petroleum ether/ethyl acetate=10/1) to afford2-(1-methylcyclopropyl)-5-nitro-1H-indole (0.70 g, 71%). ¹H NMR (400MHz, CDCl₃) δ 8.56 (brs, 1H), 8.44 (d, J=2.0 Hz, 1H), 8.01 (dd, J=2.4,8.8 Hz, 1H), 7.30 (d, J=8.8 Hz, 1H), 6.34 (d, J=1.6 Hz, 1H), 1.52 (s,3H), 1.03-0.97 (m, 2H), 0.89-0.83 (m, 2H).

2-(1-Methyl-cyclopropyl)-1H-indol-5-ylamine

To a solution of 2-(1-methylcyclopropyl)-5-nitro-1H-indole (0.70 g, 3.2mmol) in EtOH (20 mL) was added Raney Nickel (100 mg) under nitrogenatmosphere. The mixture was stirred under hydrogen atmosphere (1 atm) atroom temperature overnight. The catalyst was filtered off through acelite pad and the filtrate was evaporated under vacuum. The residue waspurified by column chromatography on silica gel (petroleum ether/ethylacetate=5/1) to afford 2-(1-methyl-cyclopropyl)-1H-indol-5-ylamine (170mg, 28%). ¹H NMR (400 MHz, CDCl₃) δ 7.65 (brs, 1H), 7.08 (d, J=8.4 Hz,1H), 6.82 (s, 1H), 6.57 (d, J=8.4 Hz, 1H), 6.14 (s, 1H), 3.45 (brs, 2H),1.47 (s, 3H), 0.82-0.78 (m, 2H), 0.68-0.63 (m, 2H).

Example 43: Methyl 2-(5-amino-1H-indol-2-yl)-2-methylpropanoate

Methyl 2,2-dimethyl-3-oxobutanoate

To a suspension of NaH (42 g, 1.1 mol, 60%) in THF (400 mL) was addeddropwise a solution of methyl 3-oxobutanoate (116 g, 1.00 mol) in THF(100 mL) at 0° C. The mixture was stirred for 0.5 h at that temperaturebefore Mel (146 g, 1.1 mol) was added dropwise at 0° C. The resultantmixture was warmed to room temperature and stirred for 1 h. NaH (42 g,1.05 mol, 60%) was added in portions at 0° C. and the resulting mixturewas continued to stir for 0.5 h at this temperature. Mel (146 g, 1.05mol) was added dropwise at 0° C. The reaction mixture was warmed to roomtemperature and stirred overnight. The mixture was poured into ice waterand the organic layer was separated. The aqueous phase was extractedwith EtOAc (500 mL×3). The combined organic layers were dried andevaporated under reduced pressure to give methyl2,2-dimethyl-3-oxobutanoate (85 g), which was used directly in the nextstep.

Methyl 3-chloro-2,2-dimethylbut-3-enoate

To a suspension of PCl₅ (270 g, 1.3 mol) in CH₂Cl₂ (1000 mL) was addeddropwise methyl 2,2-dimethyl-3-oxobutanoate (85 g) at 0° C., followingby addition of approximately 30 drops of dry DMF. The mixture was heatedat reflux overnight. The reaction mixture was cooled to ambienttemperature and slowly poured into ice water. The organic layer wasseparated and the aqueous phase was extracted with CH₂Cl₂ (500 mL×3).The combined organic layers were washed with saturated aqueous NaHCO₃and dried over anhydrous Na₂SO₄. The solvent was evaporated and theresidue was distilled under reduced pressure to give methyl3-chloro-2,2-dimethylbut-3-enoate (37 g, 23%). ¹H NMR (400 MHz, CDCl₃) δ5.33 (s, 1H), 3.73 (s, 3H), 1.44 (s, 6H).

3-Chloro-2,2-dimethylbut-3-enoic acid

A mixture of methyl 3-chloro-2,2-dimethylbut-3-enoate (33 g, 0.2 mol)and NaOH (9.6 g, 0.24 mol) in water (200 mL) was heated at reflux for 5h. The mixture was cooled to ambient temperature and extracted withether. The organic layer was discarded. The aqueous layer was acidifiedwith cold 20% HCl solution and extracted ether (200 mL×3). The combinedorganic layers were dried and evaporated under reduced pressure to give3-chloro-2,2-dimethyl-but-3-enoic acid (21 g, 70%), which was useddirectly in the next step. ¹H NMR (400 MHz, CDCl₃) δ 7.90 (brs, 1H),5.37 (dd, J=2.4, 6.8 Hz, 2H), 1.47 (s, 6H).

2,2-Dimethyl-but-3-ynoic Acid

Liquid NH₃ was condensed in a 3-neck, 250 mL round bottom flask at −78°C. Na (3.98 g, 0.173 mol) was added to the flask in portions. Themixture was stirred for 2 h at −78° C. before anhydrous DMSO (20 mL) wasadded dropwise at −78° C. The mixture was stirred at room temperatureuntil no more NH₃ was given off. A solution of3-chloro-2,2-dimethyl-but-3-enoic acid (6.5 g, 43 mmol) in DMSO (10 mL)was added dropwise at −40° C. The mixture was warmed and stirred at 50°C. for 5 h, then stirred at room temperature overnight. The cloudy,olive green solution was poured into cold 20% HCl solution and thenextracted three times with ether. The ether extracts were dried overanhydrous Na₂SO₄ and concentrated to give crude 2,2-dimethyl-but-3-ynoicacid (2 g), which was used directly in the next step. ¹H NMR (400 MHz,CDCl₃) δ 2.30 (s, 1H), 1.52 (s, 6H).

Methyl 2,2-dimethylbut-3-ynoate

To a solution of diazomethane (˜10 g) in ether (400 mL) was addeddropwise 2,2-dimethyl-but-3-ynoic acid (10.5 g, 93.7 mmol) at 0° C. Themixture was warmed to room temperature and stirred overnight. Themixture was distilled under atmospheric pressure to give crude methyl2,2-dimethylbut-3-ynoate (14 g), which was used directly in the nextstep. ¹H NMR (400 MHz, CDCl₃) δ 3.76 (s, 3H), 2.28 (s, 1H), 1.50 (s,6H).

Methyl 4-(2-amino-5-nitrophenyl)-2,2-dimethylbut-3-ynoate

To a deoxygenated solution of compound 2-bromo-4-nitroaniline (9.43 g,43.7 mmol), methyl 2,2-dimethylbut-3-ynoate (5.00 g, 39.7 mmol), CuI(754 mg, 3.97 mmol) and triethylamine (8.03 g, 79.4 mmol) in toluene/H₂O(100/30 mL) was added Pd(PPh₃)₄ (6.17 g, 3.97 mmol) under N₂. Themixture was heated at 70° C. and stirred for 24 h. After cooling, thesolid was filtered off and washed with EtOAc (50 mL×3). The organiclayer was separated and the aqueous phase was washed with EtOAc (50mL×3). The combined organic layers were dried and evaporated underreduced pressure to give a residue, which was purified by columnchromatography on silica gel (petroleum ether/ethyl acetate=10/1) toobtain methyl 4-(2-amino-5-nitrophenyl)-2,2-dimethylbut-3-ynoate (900mg, 9%). ¹H NMR (400 MHz, CDCl₃) δ 8.17 (d, J=2.8 Hz, 1H), 8.01 (dd,J=2.8, 9.2 Hz, 1H), 6.65 (d, J=9.2 Hz, 1H), 5.10 (brs, 2H), 3.80 (s,3H), 1.60 (s, 6H).

Methyl 4-(2-butyramido-5-nitrophenyl)-2,2-dimethylbut-3-ynoate

To a solution of methyl4-(2-amino-5-nitrophenyl)-2,2-dimethylbut-3-ynoate (260 mg, 1.0 mmol)and pyridine (160 mg, 2.0 mol) in CH₂Cl₂ (20 mL) was added butyrylchloride (140 mg, 1.3 mmol) at 0° C. The reaction mixture was warmed toroom temperature and stirred for 3 h before the mixture was poured intoice-water. The organic layer was separated and the aqueous phase wasextracted with CH₂Cl₂ (30 mL×3). The combined organic layers were driedover anhydrous Na₂SO₄ and evaporated under reduced pressure to obtainmethyl 4-(2-butyramido-5-nitrophenyl)-2,2-dimethylbut-3-ynoate (150 mg,45%), which was used directly in the next step. ¹H NMR (400 MHz, CDCl₃)δ 8.79 (brs, 1H), 8.71 (d, J=9.2 Hz, 1H), 8.24 (d, J=2.8 Hz, 1H), 8.17(dd, J=2.8, 9.2 Hz, 1H), 3.82 (s, 3H), 2.55 (t, J=7.2 Hz, 2H), 1.85-1.75(m, 2H), 1.63 (s, 6H), 1.06 (t, J=6.8 Hz, 3H).

Methyl 2-methyl-2-(5-nitro-1H-indol-2-yl)propanoate

To a deoxygenated solution of methyl4-(2-butyramido-5-nitrophenyl)-2,2-dimethylbut-3-ynoate (1.8 g, 5.4mmol) in acetonitrile (30 mL) was added Pd(CH₃CN)₂Cl₂ (0.42 g, 1.6=mmol)under N₂. The mixture was heated at reflux for 24 h. After cooling themixture to ambient temperature, the solid was filtered off and washedwith EtOAc (50 mL×3). The filtrate was evaporated under reduced pressureto give a residue, which was purified by column chromatography on silicagel (petroleum ether/ethyl acetate=30/1) to give methyl2-methyl-2-(5-nitro-1H-indol-2-yl)propanoate (320 mg, 23%). ¹H NMR (400MHz, CDCl₃) δ 9.05 (brs, 1H), 8.52 (d, J=2.0 Hz, 1H), 8.09 (dd, J=2.0,8.8 Hz, 1H), 7.37 (d, J=8.8 Hz, 1H), 6.54 (d, J=1.6 Hz, 1H), 3.78 (d,J=9.6 Hz, 3H), 1.70 (s, 6H).

Methyl 2-(5-amino-1H-indol-2-yl)-2-methylpropanoate

A suspension of methyl 2-methyl-2-(5-nitro-1H-indol-2-yl)propanoate (60mg, 0.23 mmol) and Raney Nickel (10 mg) in MeOH (5 mL) was hydrogenatedunder hydrogen (1 atm) at room temperature overnight. The catalyst wasfiltered off through a celite pad and the filtrate was evaporated undervacuum to give a residue, which was purified by column chromatography onsilica gel (petroleum ether/ethyl acetate=5/1) to give methyl2-(5-amino-1H-indol-2-yl)-2-methylpropanoate (20 mg, 38%). ¹H NMR (400MHz, CDCl₃) δ 8.37 (br s, 1H), 7.13 (d, J=8.4 Hz, 1H), 6.87 (d, J=2.0Hz, 1H), 6.63 (dd, J=2.0, 8.4 Hz, 1H), 6.20 (d, J=1.2 Hz, 1H), 3.72 (d,J=7.6 Hz, 3H), 3.43 (br s, 1H), 1.65 (s, 6H); MS (ESI) m/e (M+H⁺) 233.2.

Example 44: 2-Isopropyl-1H-indol-5-amine

2-Isopropyl-5-nitro-1H-indole

A mixture of methyl4-(2-butyramido-5-nitrophenyl)-2,2-dimethylbut-3-ynoate (0.50 g, 1.5mmol) and TBAF (790 mg, 3.0 mmol) in DMF (20 mL) was heated at 70° C.for 24 h. The reaction mixture was cooled to room temperature and pouredinto ice water. The mixture was extracted with ether (30 mL×3). Thecombined organic layers were dried over anhydrous Na₂SO₄ and evaporatedunder reduced pressure to give a residue, which was purified by columnchromatography on silica gel (petroleum ether/ethyl acetate=20/1) togive 2-isopropyl-5-nitro-1H-indole (100 mg, 33%). ¹H NMR (400 MHz,CDCl₃) δ 8.68 (s, 1H), 8.25 (br s, 1H), 8.21 (dd, J=2.4, 10.0 Hz, 1H),7.32 (d, J=8.8 Hz, 1H), 6.41 (s, 1H), 3.07-3.14 (m, 1H), 1.39 (d, J=6.8Hz, 6H).

2-Isopropyl-1H-indol-5-amine

A suspension of 2-isopropyl-5-nitro-1H-indole (100 mg, 0.49 mmol) andRaney Nickel (10 mg) in MeOH (10 mL) was hydrogenated under hydrogen (1atm) at the room temperature overnight. The catalyst was filtered offthrough a celite pad and the filtrate was evaporated under vacuum togive a residue, which was purified by column (petroleum ether/ethylacetate=5/1) to give 2-isopropyl-1H-indol-5-amine (35 mg, 41%). ¹H NMR(400 MHz, CDCl₃) δ 7.69 (br s, 1H), 7.10 (d, J=8.4 Hz, 1H), 6.86 (d,J=2.4 Hz, 1H), 6.58 (dd, J=2.4, 8.8 Hz, 1H), 6.07 (t, J=1.2 Hz, 1H),3.55 (br s, 2H), 3.06-2.99 (m, 1H), 1.33 (d, J=7.2 Hz, 6H); MS (ESI) m/e(M+H⁺) 175.4.

Example 45:1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-(1-hydroxy-2-methylpropan-2-yl)-1H-indol-5-yl)cyclopropanecarboxamide

Triphenyl(2-aminobenzyl)phosphonium Bromide

2-Aminobenzyl alcohol (60.0 g, 0.487 mol) was dissolved in acetonitrile(2.5 L) and brought to reflux. Triphenylphosphine hydrobromide (167 g,0.487 mol) was added and the mixture was heated at reflux for 3 h. Thereaction mixture was concentrated to approximately 500 mL and left atroom temperature for 1 h. The precipitate was filtered and washed withcold acetonitrile followed by hexane. The solid was dried overnight at40° C. under vacuum to give triphenyl(2-aminobenzyl)phosphonium bromide(193 g, 88%).

Triphenyl((ethyl(2-carbamoyl)acetate)-2-benzyl)phosphonium Bromide

To a suspension of triphenyl(2-aminobenzyl)phosphonium bromide (190 g,0.43 mol) in anhydrous dichloromethane (1 L) was added ethyl malonylchloride (55 ml, 0.43 mol). The reaction was stirred for 3 h at roomtemperature. The mixture was evaporated to dryness before ethanol (400mL) was added. The mixture was heated at reflux until a clear solutionwas obtained. The solution was left at room temperature for 3 h. Theprecipitate was filtered, washed with cold ethanol followed by hexaneand dried. A second crop was obtained from the mother liquor in the sameway. In order to remove residual ethanol both crops were combined anddissolved in dichloromethane (approximately 700 mL) under heating andevaporated. The solid was dried overnight at 50° C. under vacuum to givetriphenyl((ethyl(2-carbamoyl)acetate)-2-benzyl)-phosphonium bromide (139g, 58%).

Ethyl 2-(1H-indol-2-yl)acetate

Triphenyl((ethyl(2-carbamoyl)acetate)-2-benzyl)phosphonium bromide (32.2g, 57.3 mmol) was added to anhydrous toluene (150 mL) and the mixturewas heated at reflux. Fresh potassium tert-butoxide (7.08 g, 63.1 mmol)was added in portions over 15 minutes. Reflux was continued for another30 minutes. The mixture was filtered hot through a plug of celite andevaporated under reduced pressure. The residue was purified by columnchromatography on silica gel (0-30% ethyl acetate in hexane over 45 min)to give ethyl 2-(1H-indol-2-yl)acetate (9.12 g, 78%).

tert-Butyl 2-((ethoxycarbonyl)methyl)-1H-indole-1-carboxylate

To a solution of ethyl 2-(1H-indol-2-yl)acetate (14.7 g, 72.2 mmol) indichloromethane (150 mL) was added 4-dimethylaminopyridine (8.83 g, 72.2mmol) and di-tert-butyl carbonate (23.7 g, 108 mmol) in portions. Afterstirring for 2 h at room temperature, the mixture was diluted withdichloromethane, washed with water, dried over magnesium sulfate andpurified by silica gel chromatography (0 to 20% EtOAc in hexane) to givetert-butyl 2-((ethoxycarbonyl)methyl)-1H-indole-1-carboxylate (20.0 g,91%).

tert-Butyl 2-(2-(ethoxycarbonyl)propan-2-yl)-1H-indole-1-carboxylate

tert-Butyl 2-((ethoxycarbonyl)methyl)-1H-indole-1-carboxylate (16.7 g,54.9 mmol) was added to anhydrous THF (100 mL) and cooled to −78° C. A0.5M solution of potassium hexamethyldisilazane (165 mL, 82 mmol) wasadded slowly such that the internal temperature stayed below −60° C.Stirring was continued for 30 minutes at −78° C. To this mixture, methyliodide (5.64 mL, 91 mmol) was added. The mixture was stirred for 30 minat room temperature and then cooled to −78° C. A 0.5M solution ofpotassium hexamethyldisilazane (210 mL, 104 mmol) was added slowly andthe mixture was stirred for another 30 minutes at −78° C. More methyliodide (8.6 mL, 137 mmol) was added and the mixture was stirred for 1.5h at room temperature. The reaction was quenched with sat. aq. ammoniumchloride and partitioned between water and dichloromethane. The aqueousphase was extracted with dichloromethane and the combined organic phaseswere dried over magnesium sulfate and evaporated under reduced pressure.The residue was purified by column chromatography on silica gel (0 to20% ethylacetate in hexane) to give tert-butyl2-(2-(ethoxycarbonyl)propan-2-yl)-1H-indole-1-carboxylate (17.1 g, 94%).

Ethyl 2-(1H-indol-2-yl)-2-methylpropanoate

tert-Butyl 2-(2-(ethoxycarbonyl)propan-2-yl)-1H-indole-1-carboxylate(22.9 g, 69.1 mmol) was dissolved in dichloromethane (200 mL) before TFA(70 mL) was added. The mixture was stirred for 5 h at room temperature.The mixture was evaporated to dryness, taken up in dichloromethane andwashed with saturated sodium bicarbonate solution, water, and brine. Theproduct was purified by column chromatography on silica gel (0-20% EtOAcin hexane) to give ethyl 2-(1H-indol-2-yl)-2-methylpropanoate (12.5 g,78%).

Ethyl 2-methyl-2-(5-nitro-1H-indol-2-yl)propanoate

Ethyl 2-(1H-indol-2-yl)-2-methylpropanoate (1.0 g, 4.3 mmol) wasdissolved in concentrated sulfuric acid (6 mL) and cooled to −10° C.(salt/ice-mixture). A solution of sodium nitrate (370 mg, 4.33 mmol) inconcentrated sulfuric acid (3 mL) was added dropwise over 30 min.Stirring was continued for another 30 min at −10° C. The mixture waspoured into ice and the product was extracted with dichloromethane. Thecombined organic phases were washed with a small amount of sat. aq.sodium bicarbonate. The product was purified by column chromatography onsilica gel (5-30% EtOAc in hexane) to give ethyl2-methyl-2-(5-nitro-1H-indol-2-yl)propanoate (0.68 g, 57%).

2-Methyl-2-(5-nitro-1H-indol-2-yl)propan-1-ol

To a cooled solution of LiAlH₄ (1.0 M in THF, 1.1 mL, 1.1 mmol) in THF(5 mL) at 0° C. was added a solution of ethyl2-methyl-2-(5-nitro-1H-indol-2-yl)propanoate (0.20 g, 0.72 mmol) in THF(3.4 mL) dropwise. After addition, the mixture was allowed to warm up toroom temperature and was stirred for 3 h. The mixture was cooled to 0°C. before water (2 mL) was slowly added followed by careful addition of15% NaOH (2 mL) and water (4 mL). The mixture was stirred at roomtemperature for 0.5 h and was filtered through a short plug of celiteusing ethyl acetate. The organic layer was separated from the aqueouslayer, dried over Na₂SO₄, filtered and evaporated under reducedpressure. The residue was purified by column chromatography on silicagel (ethyl acetate/hexane=1/1) to give2-methyl-2-(5-nitro-1H-indol-2-yl)propan-1-ol (0.098 g, 58%).

2-(5-Amino-1H-indol-2-yl)-2-methylpropan-1-ol

To a solution of 2-methyl-2-(5-nitro-1H-indol-2-yl)propan-1-ol (0.094 g,0.40 mmol) in ethanol (4 mL) was added tin chloride dihydrate (0.451 g,2.0 mmol). The mixture was heated in the microwave at 120° C. for 1 h.The mixture was diluted with ethyl acetate and water before beingquenched with saturated aqueous NaHCO₃. The reaction mixture wasfiltered through a plug of celite using ethyl acetate. The organic layerwas separated from the aqueous layer, dried over Na₂SO₄, filtered andevaporated under reduced pressure to give2-(5-amino-1H-indol-2-yl)-2-methylpropan-1-ol (0.080 g, 98%).

Example 46: 2-(Pyridin-2-yl)-1H-indol-5-amine

4-Nitro-2-(pyridin-2-ylethynyl)aniline

To the solution of 2-iodo-4-nitroaniline (3.0 g, 11 mmol) in DMF (60 mL)and Et₃N (60 mL) was added 2-ethynylpyridine (3.0 g, 45 mmol),Pd(PPh₃)₂Cl₂ (600 mg) and CuI (200 mg) under N₂. The reaction mixturewas stirred at 60° C. for 12 h. The mixture was diluted with water andextracted with dichloromethane (3×100 mL). The combined organic layerswere washed with brine, dried over anhydrous Na₂SO₄ and concentrated invacuum. The residue was purified by chromatography on silica gel (5-10%ethyl acetate/petroleum ether) to afford4-nitro-2-(pyridin-2-ylethynyl)aniline (1.5 g, 60%). ¹H NMR (300 MHz,CDCl₃) δ 8.60 (s, 1H), 8.13 (d, J=2.1 Hz, 1H), 7.98 (d, J=1.8, 6.9 Hz,1H), 7.87-7.80 (m, 2H), 7.42-7.39 (m, 1H), 7.05 (brs, 2H), 6.80 (d,J=6.9 Hz, 1H).

5-Nitro-2-(pyridin-2-yl)-1H-indole

To the solution of 4-nitro-2-(pyridin-2-ylethynyl)aniline (1.5 g, 6.3mmol) in DMF (50 mL) was added t-BuOK (1.5 g, 13 mmol). The reactionmixture was stirred at 90° C. for 2 h. The mixture was diluted withwater and extracted with dichloromethane (3×50 mL). The combined organiclayers were washed with brine, dried over anhydrous Na₂SO₄ andconcentrated in vacuum. The residue was purified by chromatography onsilica gel (5-10% ethyl acetate/petroleum ether) to afford5-nitro-2-(pyridin-2-yl)-1H-indole (1.0 g, 67% yield). ¹H NMR (300 MHz,d-DMSO) δ 12.40 (s, 1H), 8.66 (d, J=2.1 Hz, 1H), 8.58 (d, J=1.8 Hz, 1H),8.07-7.91 (m, 3H), 7.59 (d, J=6.6 Hz, 1H), 7.42-7.37 (m, 2H).

2-(Pyridin-2-yl)-1H-indol-5-amine

To a solution of 5-nitro-2-(pyridin-2-yl)-1H-indole (700 mg, 2.9 mmol)in EtOH (20 mL) was added SnCl₂ (2.6 g, 12 mmol). The mixture was heatedat reflux for 10 h. Water was added and the mixture was extracted withEtOAc (50 mL×3). The combined organic layers were washed with brine,dried over anhydrous Na₂SO₄ and concentrated in vacuum. The residue waspurified by chromatography on silica gel (5-10% ethyl acetate/petroleumether) to afford 2-(pyridin-2-yl)-1H-indol-5-amine (120 mg, 20%). ¹H NMR(400 MHz, CDCl₃) δ 9.33 (brs, 1H), 8.55 (dd, J=1.2, 3.6 Hz, 1H),7.76-7.67 (m, 2 H), 7.23 (d, J=6.4 Hz, 1H), 7.16-7.12 (m, 1H), 6.94 (d,J=2.0 Hz, 1H), 6.84 (d, J=2.4 Hz, 1H), 6.71-6.69 (dd, J=2.0, 8.4 Hz,1H).

Example 47: 2-(Pyridin-2-yl)-1H-indol-5-amine

[2-(tert-Butyl-dimethyl-silanyloxy)-ethyl]-(2-iodo-4-nitro-phenyl)-amine

To a solution of 2-iodo-4-nitroaniline (2.0 g, 7.6 mmol) and2-(tert-butyldimethylsilyloxy)-acetaldehyde (3.5 g, 75% purity, 15 mmol)in methanol (30 mL) was added TFA (1.5 mL) at 0° C. The reaction mixturewas stirred at this temperature for 30 min before NaCNBH₃ (900 mg, 15mmol) was added in portions. The mixture was stirred for 2 h and wasthen quenched with water. The resulting mixture was extracted with EtOAc(30 mL×3), the combined organic extracts were dried over anhydrousNa₂SO₄ and evaporated under vacuum, and the residue was purified bychromatography on silica gel (5% ethyl acetate/petroleum) to afford[2-(tert-butyl-dimethyl-silanyloxy)-ethyl]-(2-iodo-4-nitro-phenyl)-amine(800 mg, 25%). ¹H NMR (300 MHz, CDCl₃) δ 8.57 (d, J=2.7 Hz, 1H), 8.12(dd, J=2.4, 9.0 Hz, 1H), 6.49 (d, J=9.3 Hz, 1H), 5.46 (br s, 1H), 3.89(t, J=5.4 Hz, 2H), 3.35 (q, J=5.4 Hz, 2H), 0.93 (s, 9H), 0.10 (s, 6H).

5-{2-[2-(tert-Butyl-dimethyl-silanyloxy)-ethylamino]-5-nitro-phenyl}-3,3-dimethyl-pent-4-ynoicAcid Ethyl Ester

To a solution of[2-(tert-butyl-dimethyl-silanyloxy)-ethyl]-(2-iodo-4-nitro-phenyl)-amine(800 mg, 1.9 mmol) in Et₃N (20 mL) was added Pd(PPh₃)₂Cl₂ (300 mg, 0.040mmol), CuI (76 mg, 0.040 mmol) and 3,3-dimethyl-but-1-yne (880 mg, 5.7mmol) successively under N₂ protection. The reaction mixture was heatedat 80° C. for 6 h and allowed to cool down to room temperature. Theresulting mixture was extracted with EtOAc (30 mL×3). The combinedorganic extracts were dried over anhydrous Na₂SO₄ and evaporated undervacuum to give5-{2-[2-(tert-butyl-dimethyl-silanyloxy)-ethylamino]-5-nitro-phenyl}-3,3-dimethyl-pent-4-ynoicacid ethyl ester (700 mg, 82%), which was used in the next step withoutfurther purification. ¹H NMR (400 MHz, CDCl₃) δ 8.09 (s, 1H), 8.00 (d,J=9.2 Hz, 1H), 6.54 (d, J=9.2 Hz, 1H), 6.45 (brs, 1H), 4.17-4.10 (m,4H), 3.82 (t, J=5.6 Hz, 2H), 3.43 (q, J=5.6 Hz, 2H), 2.49 (s, 2H), 1.38(s, 6H), 1.28 (t, J=7.2 Hz, 3H), 0.84 (s, 9H), 0.00 (s, 6H).

3-[1-(2-Hydroxy-ethyl)-5-nitro-1H-indol-2-yl]-3-methyl-butyric acidethyl ester

A solution of5-{2-[2-(tert-butyl-dimethyl-silanyloxy)-ethylamino]-5-nitro-phenyl}-3,3-dimethyl-pent-4-ynoicacid ethyl ester (600 mg, 1.34 mmol) and PdCl₂ (650 mg) in CH₃CN (30 mL)was heated at reflux overnight. The resulting mixture was extracted withEtOAc (30 mL×3). The combined organic extracts were dried over anhydrousNa₂SO₄ and evaporated under vacuum. The residue was dissolved in THF (20mL) and TBAF (780 mg, 3.0 mmol) was added. The mixture was stirred atroom temperature for 1 h, the solvent was removed under vacuum, and theresidue was purified by chromatography on silica gel (10% ethylacetate/petroleum) to afford3-[1-(2-hydroxy-ethyl)-5-nitro-1H-indol-2-yl]-3-methyl-butyric acidethyl ester (270 mg, 60%). ¹H NMR (300 MHz, CDCl₃) δ 8.45 (d, J=2.1 Hz,1H), 8.05 (dd, J=2.1, 9.0 Hz, 1H), 6.36 (d, J=9.0 Hz, 1H), 6.48 (s, 1H),4.46 (t, J=6.6 Hz, 2H), 4.00-3.91 (m, 4H), 2.76 (s, 2H), 1.61 (s, 6H),0.99 (t, J=7.2 Hz, 1H), 0.85 (s, 9H), 0.03 (s, 6H).

3-[1-(2-Hydroxy-ethyl)-5-nitro-1H-indol-2-yl]-3-methyl-butan-1-ol

To a solution of3-[1-(2-hydroxy-ethyl)-5-nitro-1H-indol-2-yl]-3-methyl-butyric acidethyl ester (700 mg, 2.1 mmol) in THF (25 mL) was added DIBAL-H (1.0 M,4.2 mL, 4.2 mmol) at −78° C. The mixture was stirred at room temperaturefor 1 h. Water (2 mL) was added and the resulting mixture was extractedwith EtOAc (15 mL×3). The combined organic layers were dried overanhydrous Na₂SO₄ and evaporated under vacuum. The residue was purifiedby chromatography on silica gel (15% ethyl acetate/petroleum) to afford3-[1-(2-hydroxy-ethyl)-5-nitro-1H-indol-2-yl]-3-methyl-butan-1-ol (300mg, 49%). ¹H NMR (300 MHz, d-DMSO) δ 8.42 (d, J=1.5 Hz, 1H), 7.95 (dd,J=1.2, 8.7 Hz, 1H), 6.36 (d, J=9.3 Hz, 1H), 6.50 (s, 1H), 5.25 (br s,1H), 4.46-4.42 (m, 4H), 3.69-3.66 (m, 2H), 3.24-3.21 (m, 2H), 1.42 (s,6H).

3-[5-Amino-1-(2-hydroxy-ethyl)-1H-indol-2-yl]-3-methyl-butan-1-ol

A solution of3-[1-(2-hydroxy-ethyl)-5-nitro-1H-indol-2-yl]-3-methyl-butan-1-ol (300mg, 1.03 mmol) and Raney Nickel (200 mg,) in CH₃OH (30 mL) was stirredfor 5 h at room temperature under a H₂ atmosphere. The catalyst wasfiltered through a celite pad and the filtrate was evaporated undervacuum to give a residue, which was purified by preparative TLC toafford 3-[5-amino-1-(2-hydroxy-ethyl)-1H-indol-2-yl]-3-methyl-butan-1-ol(70 mg, 26%). ¹H NMR (300 MHz, CDCl₃) δ 7.07 (d, J=8.7 Hz, 1H), 6.83 (d,J=2.1 Hz, 1H), 6.62 (dd, J=2.1, 8.4 Hz, 1H), 6.15 (s, 1H), 4.47 (t,J=5.4 Hz, 2H), 4.07 (t, J=5.4 Hz, 2H), 3.68 (t, J=5.7 Hz, 2H), 2.16 (t,J=5.7 Hz, 2H), 4.00-3.91 (m, 4H), 2.76 (s, 2H), 1.61 (s, 6H), 1.42 (s,6H).

Example 48: tert-Butyl 2-(5-amino-1H-indol-2-yl)piperidine-1-carboxylate

2-(Piperidin-2-yl)-1H-indol-5-amine

5-Nitro-2-(pyridin-2-yl)-1H-indole (1.0 g, 4.2 mmol) was added toHCl/MeOH (2 M, 50 mL). The reaction mixture was stirred at roomtemperature for 1 h and the solvent was evaporated under vacuum. PtO₂(200 mg) was added to a solution of the residue in MeOH (50 mL) and thereaction mixture was stirred under hydrogen atmosphere (1 atm) at roomtemperature for 2 h. The catalyst was filtered through a celite pad andthe solvent was evaporated under vacuum to afford2-(piperidin-2-yl)-1H-indol-5-amine (1.0 g), which was directly used inthe next step.

tert-Butyl 2-(5-amino-1H-indol-2-yl)piperidine-1-carboxylate

To a solution of 2-(piperidin-2-yl)-1H-indol-5-amine (1.0 g) in Et₃N (25mL) and THF (25 mL) was added Boc₂O (640 mg, 2.9 mmol). The reactionmixture was stirred at room temperature overnight. The mixture wasdiluted with water and extracted with dichloromethane (3×25 mL). Thecombined organic layers were washed with brine, dried over anhydrousNa₂SO₄ and concentrated in vacuum. The residue was purified bychromatography on silica gel (5-10% ethyl acetate/petroleum ether)followed by preparative HPLC to afford tert-butyl2-(5-amino-1H-indol-2-yl)piperidine-1-carboxylate (15 mg, 1% over 2steps). ¹H NMR (400 MHz, CDCl₃) δ 8.82 (s, 1H), 7.58 (s, 1H), 7.22 (d,J=8.8 Hz, 1H), 7.02 (d, J=1.6, 8.0 Hz, 1H), 6.42 (s, 1H), 6.25 (s, 1H),3.91-3.88 (m, 1H), 3.12-3.10 (m, 1H), 2.81-2.76 (m, 1H), 2.06-1.97 (m,4H), 1.70-1.58 (m, 2H), 1.53 (s, 9H).

Example 49: 6-amino-1H-indole-2-carbonitrile

(3-Nitrophenyl)hydrazine hydrochloride

3-Nitroaniline (28 g, 0.20 mol) was dissolved in a mixture of H₂O (40mL) and 37% HCl (40 mL). A solution of NaNO₂ (14 g, 0.20 mol) in H₂O (60mL) was added to the mixture at 0° C., and then a solution of SnCl₂.H₂O(140 g, 0.60 mol) in 37% HCl (100 mL) was added. After stirring at 0° C.for 0.5 h, the insoluble material was isolated by filtration and waswashed with water to give (3-nitrophenyl)hydrazine hydrochloride (28 g,73%).

(E)-Ethyl 2-(2-(3-nitrophenyl)hydrazono)propanoate

(3-Nitrophenyl)hydrazine hydrochloride (30 g, 0.16 mol) and2-oxo-propionic acid ethyl ester (22 g, 0.19 mol) were dissolved inethanol (300 mL). The mixture was stirred at room temperature for 4 hbefore the solvent was evaporated under reduced pressure to give(E)-ethyl 2-(2-(3-nitrophenyl)hydrazono)propanoate, which was useddirectly in the next step.

Ethyl 4-nitro-1H-indole-2-carboxylate and ethyl6-nitro-1H-indole-2-carboxylate

(E)-Ethyl 2-(2-(3-nitrophenyl)hydrazono)propanoate was dissolved intoluene (300 mL) and PPA (30 g) was added. The mixture was heated atreflux overnight and then was cooled to room temperature. The solventwas decanted and evaporated to obtain a crude mixture that was taken onto the next step without purification (15 g, 40%).

4-Nitro-1H-indole-2-carboxylic acid and 6-nitro-1H-indole-2-carboxylicAcid

A mixture of ethyl 6-nitro-1H-indole-2-carboxylate (0.5 g) and 10% NaOH(20 mL) was heated at reflux overnight and then was cooled to roomtemperature. The mixture was extracted with ether and the aqueous phasewas acidified with HCl to pH 1˜2. The insoluble solid was isolated byfiltration to give a crude mixture that was taken on to the next stepwithout purification (0.3 g, 68%).

4-Nitro-1H-indole-2-carboxamide and 6-nitro-1H-indole-2-carboxamide

A mixture of 6-nitro-1H-indole-2-carboxylic acid (12 g, 58 mmol) andSOCl₂ (50 mL, 64 mmol) in benzene (150 mL) was heated at reflux for 2 h.The benzene and excess SOCl₂ was removed under reduced pressure. Theresidue was dissolved in anhydrous CH₂Cl₂ (250 mL) and NH₃.H₂O (22 g,0.32 mol) was added dropwise at 0° C. The mixture was stirred at roomtemperature for 1 h. The insoluble solid was isolated by filtration toobtain crude mixture (9.0 g, 68%), which was used directly in the nextstep.

4-Nitro-1H-indole-2-carbonitrile and 6-nitro-1H-indole-2-carbonitrile

6-Nitro-1H-indole-2-carboxamide (5.0 g, 24 mmol) was dissolved in CH₂Cl₂(200 mL). Et₃N (24 g, 0.24 mol) and (CF₃CO)₂O (51 g, 0.24 mol) wereadded dropwise to the mixture at room temperature. The mixture wascontinued to stir for 1 h and was then poured into water (100 mL). Theorganic layer was separated and the aqueous layer was extracted withEtOAc (100 mL×3). The combined organic layers were dried over Na₂SO₄,filtered and concentrated under reduced pressure to obtain crude productwhich was purified by column chromatography on silica gel to give aimpure sample of 4-nitro-1H-indole-2-carbonitrile (2.5 g, 55%).

6-Amino-1H-indole-2-carbonitrile

A mixture of 6-nitro-1H-indole-2-carbonitrile (2.5 g, 13 mmol) and RaneyNickel (500 mg) in EtOH (50 mL) was stirred at room temperature under H₂(1 atm) for 1 h. Raney Nickel was removed via filtration and thefiltrate was evaporated under reduced pressure to give a residue, whichwas purified by column chromatography on silica get to give6-amino-1H-indole-2-carbonitrile (1.0 g, 49%). ¹H NMR (DMSO-d₆) δ 12.75(br s, 1H), 7.82 (d, J=8 Hz, 1H), 7.57 (s, 1H), 7.42 (s, 1H), 7.15 (d,J=8 Hz, 1H); MS (ESI) m/e (M+H⁺) 158.2.

Example 50: 6-Amino-1H-indole-3-carbonitrile

6-Nitro-1H-indole-3-carbonitrile

To a solution of 6-nitroindole (4.9 g 30 mmol) in DMF (24 mL) and CH₃CN(240 mL) was added dropwise a solution of ClSO₂NCO (5.0 mL) in CH₃CN (39mL) at 0° C. After addition, the reaction was allowed to warm to roomtemperature and was stirred for 2 h. The mixture was then poured intoice-water and basified with sat. NaHCO₃ solution to pH 7-8. The mixturewas extracted with ethyl acetate. The organics were washed with brine,dried over Na₂SO₄ and concentrated to give6-nitro-1H-indole-3-carbonitrile (4.6 g, 82%).

6-Amino-1H-indole-3-carbonitrile

A suspension of 6-nitro-1H-indole-3-carbonitrile (4.6 g, 25 mmol) and10% Pd-C (0.46 g) in EtOH (50 mL) was stirred under H₂ (1 atm) at roomtemperature overnight. After filtration, the filtrate was concentratedand the residue was purified by column chromatography on silica gel(petroleum ether/ethyl acetate=3/1) to give6-amino-1H-indole-3-carbonitrile (1.0 g, 98%) as a pink solid. ¹H NMR(DMSO-d₆) δ 11.51 (s, 1H), 7.84 (d, J=2.4 Hz, 1H), 7.22 (d, J=8.4 Hz,1H), 6.62 (s, 1H), 6.56 (d, J=8.4 Hz, 1H), 5.0 (s, 2H); MS (ESI) m/e(M+H⁺) 157.1.

Example 51: 2-tert-Butyl-1H-indol-6-amine

N-o-Tolylpivalamide

To a solution of o-tolylamine (21 g, 0.20 mol) and Et₃N (22 g, 0.22 mol)in CH₂Cl₂ was added 2,2-dimethyl-propionyl chloride (25 g, 0.21 mol) at10° C. After addition, the mixture was stirred overnight at roomtemperature. The mixture was washed with aq. HCl (5%, 80 mL), saturatedaq. NaHCO₃ and brine. The organic layer was dried over Na₂SO₄ andconcentrated under vacuum to give N-o-tolylpivalamide (35 g, 91%). ¹HNMR (300 MHz, CDCl₃) δ 7.88 (d, J=7.2 Hz, 1H), 7.15-7.25 (m, 2H), 7.05(t, J=7.2 Hz, 1H), 2.26 (s, 3H), 1.34 (s, 9H).

2-tert-Butyl-1H-indole

To a solution of N-o-tolylpivalamide (30.0 g, 159 mmol) in dry THF (100mL) was added dropwise n-BuLi (2.5 M in hexane, 190 mL) at 15° C. Afteraddition, the mixture was stirred overnight at 15° C. The mixture wascooled in an ice-water bath and treated with saturated NH₄Cl. Theorganic layer was separated and the aqueous layer was extracted withethyl acetate. The combined organic layers were dried over anhydrousNa₂SO₄, filtered, and concentrated in vacuum. The residue was purifiedby column chromatography on silica gel to give 2-tert-butyl-1H-indole(24 g, 88%). ¹H NMR (300 MHz, CDCl₃) δ 7.99 (br. s, 1H), 7.54 (d, J=7.2Hz, 1H), 7.05 (d, J=7.8 Hz, 1H), 7.06-7.13 (m, 2H), 6.26 (s, 1H), 1.39(s, 9H).

2-tert-Butylindoline

To a solution of 2-tert-butyl-1H-indole (10 g, 48 mmol) in AcOH (40 mL)was added NaBH₄ at 10° C. The mixture was stirred for 20 minutes at 10°C. before being treated dropwise with H₂O under ice cooling. The mixturewas extracted with ethyl acetate. The combined organic layers were driedover anhydrous Na₂SO₄, filtered, and concentrated under vacuum to give2-tert-butylindoline (9.8 g), which was used directly in the next step.

2-tert-butyl-6-nitroindoline and 2-tert-butyl-5-nitro-1H-indole

To a solution of 2-tert-butylindoline (9.7 g) in H₂SO₄ (98%, 80 mL) wasslowly added KNO₃ (5.6 g, 56 mmol) at 0° C. After addition, the reactionmixture was stirred at room temperature for 1 h. The mixture wascarefully poured into cracked ice, basified with Na₂CO₃ to pH 8 andextracted with ethyl acetate. The combined extracts were washed withbrine, dried over anhydrous Na₂SO₄ and concentrated under vacuum. Theresidue was purified by column chromatography to give2-tert-butyl-6-nitroindoline (4.0 g, 310% over two steps). ¹H NMR (300MHz, CDCl₃) δ 7.52 (dd, J=1.8, 8.1 Hz, 1H), 7.30 (s, 1H), 7.08 (d, J=7.8Hz, 1H), 3.76 (t, J=9.6 Hz, 1H), 2.98-3.07 (m, 1H), 2.82-2.91 (m, 1H),0.91 (s, 9H).

2-tert-Butyl-6-nitro-1H-indole

To a solution of 2-tert-butyl-6-nitroindoline (2.0 g, 9.1 mmol) in1,4-dioxane (20 mL) was added DDQ (6.9 g, 30 mmol) at room temperature.The mixture was heated at reflux for 2.5 h before being filtered andconcentrated under vacuum. The residue was purified by columnchromatography to give 2-tert-butyl-6-nitro-1H-indole (1.6 g, 80%). ¹HNMR (300 MHz, CDCl₃) δ 8.30 (br. s, 1H), 8.29 (s, 1H), 8.00 (dd, J=2.1,8.7 Hz, 1H), 7.53 (d, J=9.3 Hz, 1H), 6.38 (s, 1H), 1.43 (s, 9H).

2-tert-Butyl-1H-indol-6-amine

To a solution of 2-tert-butyl-6-nitro-1H-indole (1.3 g, 6.0 mmol) inMeOH (10 mL) was added Raney Nickel (0.2 g). The mixture washydrogenated under 1 atm of hydrogen at room temperature for 3 h. Thereaction mixture was filtered and the filtrate was concentrated. Theresidue was washed with petroleum ether to give2-tert-butyl-1H-indol-6-amine (1.0 g, 89%). ¹H NMR (300 MHz, DMSO-d₆) δ10.19 (s, 1H), 6.99 (d, J=8.1 Hz, 1H), 6.46 (s, 1H), 6.25 (dd, J=1.8,8.1 Hz, 1H), 5.79 (d, J=1.8 Hz, 1H), 4.52 (s, 2H), 1.24 (s, 9H); MS(ESI) m/e (M+H⁺) 189.1.

Example 52: 3-tert-Butyl-1H-indol-6-amine

3-tert-Butyl-6-nitro-1H-indole

To a mixture of 6-nitroindole (1.0 g, 6.2 mmol), zinc triflate (2.1 g,5.7 mmol), and TBAI (1.7 g, 5.2 mmol) in anhydrous toluene (11 mL) wasadded DIEA (1.5 g, 11 mmol) at room temperature under nitrogen. Thereaction mixture was stirred for 10 min at 120° C., followed by theaddition of t-butyl bromide (0.71 g, 5.2 mmol). The resulting mixturewas stirred for 45 min at 120° C. The solid was filtered off and thefiltrate was concentrated to dryness. The residue was purified by columnchromatography on silica gel (petroleum ether/ethyl acetate=20:1) togive 3-tert-butyl-6-nitro-1H-indole (0.25 g, 19%) as a yellow solid.¹H-NMR (CDCl₃) δ 8.32 (d, J=2.1 Hz, 1H), 8.00 (dd, J=2.1, 14.4 Hz, 1H),7.85 (d, J=8.7 Hz, 1H), 7.25 (s, 1H), 1.46 (s, 9H).

3-tert-Butyl-1H-indol-6-amine

A suspension of 3-tert-butyl-6-nitro-1H-indole (3.0 g, 14 mmol) andRaney Nickel (0.5 g) was hydrogenated under H₂ (1 atm) at roomtemperature for 3 h. The catalyst was filtered off and the filtrate wasconcentrated to dryness. The residue was purified by column on silicagel (petroleum ether/ethyl acetate=4:1) to give3-tert-butyl-1H-indol-6-amine (2.0 g, 77%) as a gray solid. ¹HNMR(CDCl₃) δ 7.58 (m, 2H), 6.73 (d, J=1.2 Hz, 1H), 6.66 (s, 1H), 6.57 (dd,J=0.8, 8.6 Hz, 1H), 3.60 (br, 2H), 1.42 (s, 9H).

Example 53: 5-(Trifluoromethyl)-1H-indol-6-amine

1-Methyl-2,4-dinitro-5-(trifluoromethyl)benzene

To a mixture of HNO₃ (98%, 30 mL) and H₂SO₄ (98%, 30 mL) was addeddropwise 1-methyl-3-trifluoromethyl-benzene (10 g, 63 mmol) at 0° C.After addition, the mixture was stirred at rt for 30 min and was thenpoured into ice-water. The precipitate was filtered and washed withwater to give 1-methyl-2,4-dinitro-5-trifluoromethyl-benzene (2.0 g,13%).

(E)-2-(2,4-Dinitro-5-(trifluoromethyl)phenyl)-N,N-dimethylethenamine

A mixture of 1-methyl-2,4-dinitro-5-trifluoromethyl-benzene (2.0 g, 8.0mmol) and DMA (1.0 g, 8.2 mmol) in DMF (20 mL) was stirred at 100° C.for 30 min. The mixture was poured into ice-water and stirred for 1 h.The precipitate was filtered and washed with water to give(E)-2-(2,4-dinitro-5-(trifluoromethyl)phenyl)-N,N-dimethylethenamine(2.1 g, 86%).

5-(Trifluoromethyl)-1H-indol-6-amine

A suspension of(E)-2-(2,4-dinitro-5-(trifluoromethyl)phenyl)-N,N-dimethylethenamine(2.1 g, 6.9 mmol) and Raney Nickel (1 g) in ethanol (80 mL) was stirredunder H₂ (1 atm) at room temperature for 5 h. The catalyst was filteredoff and the filtrate was concentrated to dryness. The residue waspurified by column on silica gel to give5-(trifluoromethyl)-1H-indol-6-amine (200 mg, 14%). ¹H NMR (DMSO-d₆) δ10.79 (br s, 1H), 7.55 (s, 1H), 7.12 (s, 1H), 6.78 (s, 1H), 6.27 (s,1H), 4.92 (s, 2H); MS (ESI) m/e (M+H⁺): 200.8.

Example 54: 5-Ethyl-1H-indol-6-amine

1-(Phenylsulfonyl)indoline

To a mixture of DMAP (1.5 g), benzenesulfonyl chloride (24.0 g, 136mmol) and indoline (14.7 g, 124 mmol) in CH₂Cl₂ (200 mL) was addeddropwise Et₃N (19.0 g, 186 mmol) at 0° C. The mixture was stirred atroom temperature overnight. The organic layer was washed with water(2×), dried over Na₂SO₄ and concentrated to dryness under reducedpressure to obtain 1-(phenylsulfonyl)indoline (30.9 g, 96%).

1-(1-(Phenylsulfonyl)indolin-5-yl)ethanone

To a suspension of AlCl₃ (144 g, 1.08 mol) in CH₂Cl₂ (1070 mL) was addedacetic anhydride (54 mL). The mixture was stirred for 15 minutes beforea solution of 1-(phenylsulfonyl)indoline (46.9 g, 0.180 mol) in CH₂Cl₂(1070 mL) was added dropwise. The mixture was stirred for 5 h and wasquenched by the slow addition of crushed ice. The organic layer wasseparated and the aqueous layer was extracted with CH₂Cl₂. The combinedorganics were washed with saturated aqueous NaHCO₃ and brine, dried overNa₂SO₄, and concentrated under vacuum to obtain1-(1-(phenylsulfonyl)indolin-5-yl)ethanone (42.6 g).

5-Ethyl-1-(phenylsulfonyl)indoline

To TFA (1600 mL) at 0° C. was added sodium borohydride (64.0 g, 1.69mol) over 1 h. To this mixture was added dropwise a solution of1-(1-(phenylsulfonyl)indolin-5-yl)ethanone (40.0 g, 0.133 mol) in TFA(700 mL) over 1 h. The mixture was then stirred overnight at 25° C.After dilution with H₂O (1600 mL), the mixture was made basic by theaddition of sodium hydroxide pellets at 0° C. The organic layer wasseparated and the aqueous layer was extracted with CH₂Cl₂. The combinedorganic layers were washed with brine, dried over Na₂SO₄ andconcentrated under reduced pressure. The residue was purified by silicacolumn to give 5-ethyl-1-(phenylsulfonyl)indoline (16.2 g, 47% over twosteps).

5-Ethylindoline

A mixture of 5-ethyl-1-(phenylsulfonyl)indoline (15 g, 0.050 mol) in HBr(48%, 162 mL) was heated at reflux for 6 h. The mixture was basifiedwith sat. NaOH to pH 9 and then it was extracted with ethyl acetate. Theorganic layer was washed with brine, dried over Na₂SO₄, and concentratedunder reduced pressure. The residue was purified by silica column togive 5-ethylindoline (2.5 g, 32%).

5-Ethyl-6-nitroindoline

To a solution of 5-ethylindoline (2.5 g, 17 mmol) in H₂SO₄ (98%, 20 mL)was slowly added KNO₃ (1.7 g, 17 mmol) at 0° C. The mixture was stirredat 0-10° C. for 10 minutes. The mixture was then carefully poured intoice, basified with NaOH solution to pH 9, and extracted with ethylacetate. The combined extracts were washed with brine, dried over Na₂SO₄and concentrated to dryness. The residue was purified by silica columnto give 5-ethyl-6-nitroindoline (1.9 g, 58%).

5-Ethyl-6-nitro-1H-indole

To a solution of 5-ethyl-6-nitroindoline (1.9 g, 9.9 mmol) in CH₂Cl₂ (30mL) was added MnO₂ (4.0 g, 46 mmol). The mixture was stirred at ambienttemperature for 8 h. The solid was filtered off and the filtrate wasconcentrated to dryness to give 5-ethyl-6-nitro-TH-indole (1.9 g).

5-Ethyl-1H-indol-6-amine

A suspension of 5-ethyl-6-nitro-1H-indole (1.9 g, 10 mmol) and RaneyNickel (1 g) was hydrogenated under H₂ (1 atm) at room temperature for 2h. The catalyst was filtered off and the filtrate was concentrated todryness. The residue was purified by silica gel column to give5-ethyl-TH-indol-6-amine (760 mg, 48% over two steps). ¹H NMR (CDCl₃) δ7.90 (br s, 1H), 7.41 (s, 1H), 7.00 (s, 1H), 6.78 (s, 2H), 6.39 (s, 1H),3.39 (br s, 2H), 2.63 (q, J=7.2 Hz, 2H), 1.29 (t, J=6.9 Hz, 3H); MS(ESI) m/e (M+H⁺) 161.1.

Example 55: Ethyl 6-amino-1H-indole-4-carboxylate

2-Methyl-3,5-dinitrobenzoic Acid

To a mixture of HNO₃ (95%, 80 mL) and H₂SO₄ (98%, 80 mL) was slowlyadded 2-methylbenzic acid (50 g, 0.37 mol) at 0° C. After addition, thereaction mixture was stirred below 30° C. for 1.5 h. The mixture thenwas poured into ice-water and stirred for 15 min. The precipitate wasfiltered and washed with water to give 2-methyl-3,5-dinitrobenzoic acid(70 g, 84%).

Ethyl 2-methyl-3,5-dinitrobenzoate

A mixture of 2-methyl-3,5-dinitrobenzoic acid (50 g, 0.22 mol) in SOCl₂(80 mL) was heated at reflux for 4 h and then was concentrated todryness. The residue was dissolved in CH₂Cl₂ (50 mL), to which EtOH (80mL) was added and the mixture was stirred at room temperature for 1 h.The mixture was poured into ice-water and extracted with EtOAc (3×100mL). The combined extracts were washed sat. Na₂CO₃ (80 mL), water (2×100mL) and brine (100 mL), dried over Na₂SO₄ and concentrated to dryness togive ethyl 2-methyl-3,5-dinitrobenzoate (50 g, 88%)

(E)-Ethyl 2-(2-(dimethylamino)vinyl)-3,5-dinitrobenzoate

A mixture of ethyl 2-methyl-3,5-dinitrobenzoate (35 g, 0.14 mol) and DMA(32 g, 0.27 mol) in DMF (200 mL) was heated at 100° C. for 5 h. Themixture was poured into ice-water and the precipitated solid wasfiltered and washed with water to give (E)-ethyl2-(2-(dimethylamino)vinyl)-3,5-dinitrobenzoate (11 g, 48%)

Ethyl 6-amino-1H-indole-4-carboxylate

A mixture of (E)-ethyl 2-(2-(dimethylamino)vinyl)-3,5-dinitrobenzoate(11 g, 0.037 mol) and SnCl₂ (83 g, 0.37 mol) in ethanol was heated atreflux for 4 h. The mixture was concentrated to dryness and the residuewas poured into water and basified using sat. aq. Na₂CO₃ to pH 8. Theprecipitated solid was filtered and the filtrate was extracted withethyl acetate (3×100 mL). The combined extracts were washed with water(2×100 mL) and brine (150 mL), dried over Na₂SO₄, and concentrated todryness. The residue was purified by column on silica gel to give ethyl6-amino-1H-indole-4-carboxylate (3.0 g, 40%). ¹HNMR (DMSO-d₆) δ 10.76(br s, 1H), 7.11-7.14 (m, 2H), 6.81-6.82 (m, 1H), 6.67-6.68 (m, 1H),4.94 (br s, 2H), 4.32-4.25 (q, J=7.2 Hz, 2H), 1.35-1.31 (t, J=7.2, 3H);MS (ESI) m/e (M+H⁺) 205.0.

Example 56: 5-Fluoro-1H-indol-6-amine

1-Fluoro-5-methyl-2,4-dinitrobenzene

To a stirred solution of HNO₃ (60 mL) and H₂SO₄ (80 mL) was addeddropwise 1-fluoro-3-methylbenzene (28 g, 25 mmol) under ice-cooling atsuch a rate that the temperature did not rise above 35° C. The mixturewas allowed to stir for 30 min at rt and was then poured into ice water(500 mL). The resulting precipitate (a mixture of1-fluoro-5-methyl-2,4-dinitrobenzene and1-fluoro-3-methyl-2,4-dinitrobenzene, 32 g, ca. 7:3 ratio) was collectedby filtration and purified by recrystallization from 50 mL isopropylether to give pure 1-fluoro-5-methyl-2,4-dinitro-benzene as a whitesolid (18 g, 36%).

(E)-2-(5-Fluoro-2,4-dinitrophenyl)-N,N-dimethylethenamine

A mixture of 1-fluoro-5-methyl-2,4-dinitro-benzene (10 g, 50 mmol), DMA(12 g, 100 mmol) and DMF (50 mL) was heated at 100° C. for 4 h. Thesolution was cooled and poured into water. The precipitated red solidwas collected, washed with water, and dried to give(E)-2-(5-fluoro-2,4-dinitrophenyl)-N,N-dimethylethenamine (8.0 g, 63%).

5-Fluoro-1H-indol-6-amine

A suspension of(E)-2-(5-fluoro-2,4-dinitrophenyl)-N,N-dimethylethenamine (8.0 g, 31mmol) and Raney Nickel (8 g) in EtOH (80 mL) was stirred under H₂ (40psi) at room temperature for 1 h. After filtration, the filtrate wasconcentrated and the residue was purified by column chromatography(petroleum ether/ethyl acetate=5/1) to give 5-fluoro-1H-indol-6-amine(1.0 g, 16%) as a brown solid. ¹HNMR (DMSO-d₆) δ 10.56 (br s, 1H), 7.07(d, J=12 Hz, 1H), 7.02 (m, 1H), 6.71 (d, J=8 Hz, 1H), 6.17 (s, 1H), 3.91(br s, 2H); MS (ESI) m/e (M+H⁺) 150.1.

Example 57: 5-Chloro-1H-indol-6-amine

1-Chloro-5-methyl-2,4-dinitrobenzene

To a stirred solution of HNO₃ (55 mL) and H₂SO₄ (79 mL) was addeddropwise 1-chloro-3-methylbenzene (25.3 g, 200 mmol) under ice-coolingat such a rate that the temperature did not rise above 35° C. Themixture was allowed to stir for 30 min at ambient temperature and wasthen poured into ice water (500 mL). The resulting precipitate wascollected by filtration and purified by recrystallization to give1-chloro-5-methyl-2,4-dinitrobenzene (26 g, 60%).

(E)-2-(5-Chloro-2,4-dinitrophenyl)-N,N-dimethylethenamine

A mixture of 1-chloro-5-methyl-2,4-dinitro-benzene (11.6 g, 50.0 mmol),DMA (11.9 g, 100 mmol) in DMF (50 mL) was heated at 100° C. for 4 h. Thesolution was cooled and poured into water. The precipitated red solidwas collected by filtration, washed with water, and dried to give(E)-2-(5-chloro-2,4-dinitrophenyl)-N,N-dimethylethenamine (9.84 g, 72%).

5-Chloro-1H-indol-6-amine

A suspension of(E)-2-(5-chloro-2,4-dinitrophenyl)-N,N-dimethylethenamine (9.8 g, 36mmol) and Raney Nickel (9.8 g) in EtOH (140 mL) was stirred under H₂ (1atm) at room temperature for 4 h. After filtration, the filtrate wasconcentrated and the residue was purified by column chromatograph(petroleum ether/ethyl acetate=10:1) to give 5-chloro-1H-indol-6-amine(0.97 g, 16%) as a gray powder. ¹HNMR (CDCl₃) δ 7.85 (br s, 1H), 7.52(s, 1H), 7.03 (s, 1H), 6.79 (s, 1H), 6.34 (s, 1H), 3.91 (br s, 1H); MS(ESI) m/e (M+H⁺) 166.0.

Example 58: Ethyl 6-amino-1H-indole-7-carboxylate

3-Methyl-2,6-dinitrobenzoic Acid

To a mixture of HNO₃ (95%, 80 mL) and H₂SO₄ (98%, 80 mL) was slowlyadded 3-methylbenzic acid (50 g, 0.37 mol) at 0° C. After addition, themixture was stirred below 30° C. for 1.5 hours. The mixture was thenpoured into ice-water and stirred for 15 min. The precipitate solid wasfiltered and washed with water to give a mixture of3-methyl-2,6-dinitro-benzoic acid and 5-methyl-2,4-dinitrobenzoic acid(70 g, 84%). To a solution of this mixture (70 g, 0.31 mol) in EtOH (150mL) was added dropwise SOCl₂ (54 g, 0.45 mol). The mixture was heated atreflux for 2 h before being concentrated to dryness under reducedpressure. The residue was partitioned between EtOAc (100 mL) and aq.Na₂CO₃ (10%, 120 mL). The organic layer was washed with brine (50 mL),dried over Na₂SO₄, and concentrated to dryness to obtain ethyl5-methyl-2,4-dinitrobenzoate (20 g), which was placed aside. The aqueouslayer was acidified by HCl to pH 2˜3 and the precipitated solid wasfiltered, washed with water, and dried in air to give3-methyl-2,6-dinitrobenzoic acid (39 g, 47%).

Ethyl 3-methyl-2,6-dinitrobenzoate

A mixture of 3-methyl-2,6-dinitrobenzoic acid (39 g, 0.15 mol) and SOCl₂(80 mL) was heated at reflux 4 h. The excess SOCl₂ was evaporated offunder reduced pressure and the residue was added dropwise to a solutionof EtOH (100 mL) and Et₃N (50 mL). The mixture was stirred at 20° C. for1 h and then concentrated to dryness. The residue was dissolved in EtOAc(100 mL), washed with Na₂CO₃ (10%, 40 mL×2), water (50 mL×2) and brine(50 mL), dried over Na₂SO₄ and concentrated to give ethyl3-methyl-2,6-dinitrobenzoate (20 g, 53%).

(E)-Ethyl 3-(2-(dimethylamino)vinyl)-2,6-dinitrobenzoate

A mixture of ethyl 3-methyl-2,6-dinitrobenzoate (35 g, 0.14 mol) and DMA(32 g, 0.27 mol) in DMF (200 mL) was heated at 100° C. for 5 h. Themixture was poured into ice water. The precipitated solid was filteredand washed with water to give (E)-ethyl3-(2-(dimethylamino)vinyl)-2,6-dinitrobenzoate (25 g, 58%).

Ethyl 6-amino-1H-indole-7-carboxylate

A mixture of (E)-ethyl 3-(2-(dimethylamino)vinyl)-2,6-dinitrobenzoate(30 g, 0.097 mol) and Raney Nickel (10 g) in EtOH (1000 mL) washydrogenated at room temperature under 50 psi for 2 h. The catalyst wasfiltered off and the filtrate was concentrated to dryness. The residuewas purified by column on silica gel to give ethyl6-amino-1H-indole-7-carboxylate as an off-white solid (3.2 g, 16%). ¹HNMR (DMSO-d₆) δ 10.38 (s, 1H), 7.42 (d, J=8.7 Hz, 1H), 6.98 (t, J=3.0Hz, 1H), 6.65 (s, 2H), 6.48 (d, J=8.7 Hz, 1H), 6.27-6.26 (m, 1H), 4.38(q, J=7.2 Hz, 2H), 1.35 (t, J=7.2 Hz, 3H).

Example 59: Ethyl 6-amino-1H-indole-5-carboxylate

(E)-Ethyl 5-(2-(dimethylamino)vinyl)-2,4-dinitrobenzoate

A mixture of ethyl 5-methyl-2,4-dinitrobenzoate (39 g, 0.15 mol) and DMA(32 g, 0.27 mol) in DMF (200 mL) was heated at 100° C. for 5 h. Themixture was poured into ice water and the precipitated solid wasfiltered and washed with water to afford (E)-ethyl5-(2-(dimethylamino)vinyl)-2,4-dinitrobenzoate (15 g, 28%).

Ethyl 6-amino-1H-indole-5-carboxylate

A mixture of (E)-ethyl 5-(2-(dimethylamino)vinyl)-2,4-dinitrobenzoate(15 g, 0.050 mol) and Raney Nickel (5 g) in EtOH (500 mL) washydrogenated at room temperature under 50 psi of hydrogen for 2 h. Thecatalyst was filtered off and the filtrate was concentrated to dryness.The residue was purified by column on silica gel to give ethyl6-amino-1H-indole-5-carboxylate (3.0 g, 30%). ¹H NMR (DMSO-d₆) δ 10.68(s, 1H), 7.99 (s, 1H), 7.01-7.06 (m, 1H), 6.62 (s, 1H), 6.27-6.28 (m,1H), 6.16 (s, 2H), 4.22 (q, J=7.2 Hz, 2H), 1.32-1.27 (t, J=7.2 Hz, 3H).

Example 60: 5-tert-Butyl-1H-indol-6-amine

2-tert-Butyl-4-methylphenyl Diethyl Phosphate

To a suspension of NaH (60% in mineral oil, 8.4 g, 0.21 mol) in THF (200mL) was added dropwise a solution of 2-tert-butyl-4-methylphenol (33 g,0.20 mol) in THF (100 mL) at 0° C. The mixture was stirred at 0° C. for15 min and then phosphorochloridic acid diethyl ester (37 g, 0.21 mol)was added dropwise at 0° C. After addition, the mixture was stirred atambient temperature for 30 min. The reaction was quenched with sat.NH₄Cl (300 mL) and then extracted with Et₂O (350 mL×2). The combinedorganic layers were washed with brine, dried over anhydrous Na₂SO₄, andthen evaporated under vacuum to give 2-tert-butyl-4-methylphenyl diethylphosphate (contaminated with mineral oil) as a colorless oil (60 g,˜100%), which was used directly in the next step.

1-tert-Butyl-3-methylbenzene

To NH₃ (liquid, 1000 mL) was added a solution of2-tert-butyl-4-methylphenyl diethyl phosphate (60 g, crude from laststep, about 0.2 mol) in Et₂O (anhydrous, 500 mL) at −78° C. under N₂atmosphere. Lithium metal was added to the solution in small piecesuntil the blue color persisted. The reaction mixture was stirred at −78°C. for 15 min and then was quenched with sat. NH₄Cl until the mixtureturned colorless. Liquid NH₃ was evaporated and the residue wasdissolved in water. The mixture was extracted with Et₂O (400 mL×2). Thecombined organics were dried over Na₂SO₄ and evaporated to give1-tert-butyl-3-methylbenzene (contaminated with mineral oil) as acolorless oil (27 g, 91%), which was used directly in next step.

1-tert-Butyl-5-methyl-2,4-dinitrobenzene and1-tert-butyl-3-methyl-2,4-dinitro-benzene

To HNO₃ (95%, 14 mL) was added H₂SO₄ (98%, 20 mL) at 0° C. and then1-tert-butyl-3-methylbenzene (7.4 g, ˜50 mmol, crude from last step)dropwise to the with the temperature being kept below 30° C. The mixturewas stirred at ambient temperature for 30 min, poured onto crushed ice(100 g), and extracted with EtOAc (50 mL×3). The combined organic layerswere washed with water and brine, before being evaporated to give abrown oil, which was purified by column chromatography to give a mixtureof 1-tert-butyl-5-methyl-2,4-dinitrobenzene and1-tert-butyl-3-methyl-2,4-dinitrobenzene (2:1 by NMR) as a yellow oil(9.0 g, 61%).

(E)-2-(5-tert-Butyl-2,4-dinitrophenyl)-N,N-dimethylethenamine

A mixture of 1-tert-butyl-5-methyl-2,4-dinitrobenzene and1-tert-butyl-3-methyl-2,4-dinitrobenzene (9.0 g, 38 mmol, 2:1 by NMR)and DMA (5.4 g, 45 mmol) in DMF (50 mL) was heated at reflux for 2 hbefore being cooled to room temperature. The reaction mixture was pouredinto water-ice and extracted with EtOAc (50 mL×3). The combined organiclayers were washed with water and brine, before being evaporated to givea brown oil, which was purified by column to give(E)-2-(5-tert-butyl-2,4-dinitrophenyl)-N,N-dimethylethen-amine (5.0 g,68%).

5-tert-Butyl-1H-indol-6-amine

A solution of(E)-2-(5-tert-butyl-2,4-dinitrophenyl)-N,N-dimethylethen-amine (5.3 g,18 mmol) and tin (II) chloride dihydrate (37 g, 0.18 mol) in ethanol(200 mL) was heated at reflux overnight. The mixture was cooled to roomtemperature and the solvent was removed under vacuum. The residualslurry was diluted with water (500 mL) and was basified with 10% aq.Na₂CO₃ to pH 8. The resulting suspension was extracted with ethylacetate (3×100 mL). The ethyl acetate extract was washed with water andbrine, dried over Na₂SO₄, and concentrated. The residual solid waswashed with CH₂Cl₂ to afford a yellow powder, which was purified bycolumn chromatography to give 5-tert-butyl-1H-indol-6-amine (0.40 g,12%). ¹H NMR (DMSO-d₆) δ 10.34 (br s, 1H), 7.23 (s, 1H), 6.92 (s, 1H),6.65 (s, 1H), 6.14 (s, 1H), 4.43 (br s, 2H), 2.48 (s, 9H); MS (ESI) m/e(M+H⁺) 189.1.

General Procedure IV: Synthesis of Acylaminoindoles

One equivalent of the appropriate carboxylic acid and one equivalent ofthe appropriate amine were dissolved in N,N-dimethylformamide (DMF)containing triethylamine (3 equivalents).O-(7-Azabenzotriazol-1-yl)-N,N,N′,N′-tetramethyluroniumhexafluorophosphate (HATU) was added and the solution was allowed tostir. The crude product was purified by reverse-phase preparative liquidchromatography to yield the pure product.

Example 61:N-(2-tert-Butyl-1H-indol-5-yl)-1-(4-methoxyphenyl)-cyclopropanecarboxamide

2-tert-Butyl-1H-indol-5-amine (19 mg, 0.10 mmol) and1-(4-methoxyphenyl)-cyclopropanecarboxylic acid (19 mg, 0.10 mmol) weredissolved in N,N-dimethylformamide (1.00 mL) containing triethylamine(28 μL, 0.20 mmol).O-(7-Azabenzotriazol-1-yl)-N,N,N′,N′-tetramethyluroniumhexafluorophosphate (42 mg, 0.11 mmol) was added to the mixture and theresulting solution was allowed to stir for 3 hours. The crude reactionmixture was filtered and purified by reverse phase HPLC. ESI-MS m/zcalc. 362.2, found 363.3 (M+1)⁺; Retention time 3.48 minutes.

General Procedure V: Synthesis of Acylaminoindoles

One equivalent of the appropriate carboxylic acid was placed in anoven-dried flask under nitrogen. A minimum (3 equivalents) of thionylchloride and a catalytic amount of and N,N-dimethylformamide were addedand the solution was allowed to stir for 20 minutes at 60° C. The excessthionyl chloride was removed under vacuum and the resulting solid wassuspended in a minimum of anhydrous pyridine. This solution was slowlyadded to a stirred solution of one equivalent the appropriate aminedissolved in a minimum of anhydrous pyridine. The resulting mixture wasallowed to stir for 15 hours at 110° C. The mixture was evaporated todryness, suspended in dichloromethane, and then extracted three timeswith 1N HCl. The organic layer was then dried over sodium sulfate,evaporated to dryness, and then purified by column chromatography.

Example 62: Ethyl5-(1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1H-indole-2-carboxylate(Compd. 28)

1-Benzo[1,3]dioxol-5-yl-cyclopropanecarboxylic acid (2.07 g, 10.0 mmol)was dissolved in thionyl chloride (2.2 mL) under N₂.N,N-dimethylformamide (0.3 mL) was added and the solution was allowed tostir for 30 minutes. The excess thionyl chloride was removed undervacuum and the resulting solid was dissolved in anhydrousdichloromethane (15 mL) containing triethylamine (2.8 mL, 20.0 mmol).Ethyl 5-amino-1H-indole-2-carboxylate (2.04 g, 10.0 mmol) in 15 mL ofanhydrous dichloromethane was slowly added to the reaction. Theresulting solution was allowed to stir for 1 hour. The reaction mixturewas diluted to 50 mL with dichloromethane and washed three times with 50mL of 1N HCl, saturated aqueous sodium bicarbonate, and saturatedaqueous sodium chloride. The organic layer was dried over sodium sulfateand evaporated to dryness to yield ethyl5-(1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1H-indole-2-carboxylateas a gray solid (3.44 g, 88%). ESI-MS m/z calc. 392.4; found 393.1(M+1)⁺ Retention time 3.17 minutes. ¹H NMR (400 MHz, DMSO-d6) δ 11.80(s, 1H), 8.64 (s, 1H), 7.83 (m, 1H), 7.33-7.26 (m, 2H), 7.07 (m, 1H),7.02 (m, 1H), 6.96-6.89 (m, 2H), 6.02 (s, 2H), 4.33 (q, J=7.1 Hz, 2H),1.42-1.39 (m, 2H), 1.33 (t, J=7.1 Hz, 3H), 1.06-1.03 (m, 2H).

Example 63:1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-1H-indol-5-yl)cyclopropanecarboxamide

1-Benzo[1,3]dioxol-5-yl-cyclopropanecarboxylic acid (1.09 g, 5.30 mmol)was dissolved in 2 mL of thionyl chloride under nitrogen. A catalyticamount (0.3 mL) of N,N-dimethylformamide (DMF) was added and thereaction mixture was stirred for 30 minutes. The excess thionyl chloridewas evaporated and the resulting residue was dissolved in 15 mL ofdichloromethane. This solution was slowly added to a solution of2-tert-butyl-1H-indol-5-amine (1.0 g, 5.3 mmol) in 10 mL ofdichloromethane containing triethylamine (1.69 mL, 12.1 mmol). Theresulting solution was allowed to stir for 10 minutes. The solvent wasevaporated to dryness and the crude reaction mixture was purified bysilica gel column chromatography using a gradient of 5-50% ethyl acetatein hexanes. The pure fractions were combined and evaporated to drynessto yield a pale pink powder (1.24 g 62%). ESI-MS m z calc. 376.18, found377.3 (M+1)⁺. Retention time of 3.47 minutes. ¹H NMR (400 MHz, DMSO) δ10.77 (s, 1H), 8.39 (s, 1H), 7.56 (d, J=1.4 Hz, 1H), 7.15 (d, J=8.6 Hz,1H), 7.05-6.87 (m, 4H), 6.03 (s, 3H), 1.44-1.37 (m, 2H), 1.33 (s, 9H),1.05-1.00 (m, 2H).

Example 64:1-(Benzo[d][1,3]dioxol-5-yl)-N-(1-methyl-2-(1-methylcyclopropyl)-1H-indol-5-yl)cyclopropanecarboxamide

1-Methyl-2-(1-methylcyclopropyl)-1H-indol-5-amine (20.0 mg, 0.100 mmol)and 1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxylic acid (20.6 mg,0.100 mmol) were dissolved in N,N-dimethylformamide (1 mL) containingtriethylamine (42.1 μL, 0.300 mmol) and a magnetic stir bar.O-(7-Azabenzotriazol-1-yl)-N,N,N′,N′-tetramethyluroniumhexafluorophosphate (42 mg, 0.11 mmol) was added to the mixture and theresulting solution was allowed to stir for 6 h at 80° C. The crudeproduct was then purified by preparative HPLC utilizing a gradient of0-99% acetonitrile in water containing 0.05% trifluoroacetic acid toyield1-(benzo[d][1,3]dioxol-5-yl)-N-(1-methyl-2-(1-methylcyclopropyl)-1H-indol-5-yl)cyclopropanecarboxamide.ESI-MS m/z calc. 388.2, found 389.2 (M+1)⁺. Retention time of 3.05minutes.

Example 65:1-(Benzo[d][1,3]dioxol-5-yl)-N-(1,1-dimethyl-2,3-dihydro-1H-pyrrolo[1,2-a]indol-7-yl)cyclopropanecarboxamide

1,1-Dimethyl-2,3-dihydro-1H-pyrrolo[1,2-a]indol-7-amine (40.0 mg, 0.200mmol) and 1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxylic acid (41.2mg, 0.200 mmol) were dissolved in N,N-dimethylformamide (1 mL)containing triethylamine (84.2 μL, 0.600 mmol) and a magnetic stir bar.O-(7-Azabenzotriazol-1-yl)-N,N,N′,N′-tetramethyluroniumhexafluorophosphate (84 mg, 0.22 mmol) was added to the mixture and theresulting solution was allowed to stir for 5 minutes at roomtemperature. The crude product was then purified by preparative HPLCutilizing a gradient of 0-99% acetonitrile in water containing 0.05%trifluoroacetic acid to yield1-(benzo[d][1,3]dioxol-5-yl)-N-(1,1-dimethyl-2,3-dihydro-1H-pyrrolo[1,2-a]-indol-7-yl)cyclopropanecarboxamide.ESI-MS m/z calc. 388.2, found 389.2 (M+1)⁺. Retention time of 2.02minutes. ¹H NMR (400 MHz, DMSO-d6) δ 8.41 (s, 1H), 7.59 (d, J=1.8 Hz,1H), 7.15 (d, J=8.6 Hz, 1H), 7.06-7.02 (m, 2H), 6.96-6.90 (m, 2H), 6.03(s, 2H), 5.98 (d, J=0.7 Hz, 1H), 4.06 (t, J=6.8 Hz, 2H), 2.35 (t, J=6.8Hz, 2H), 1.42-1.38 (m, 2H), 1.34 (s, 6H), 1.05-1.01 (m, 2H).

Example 66: Methyl5-(1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-2-tert-butyl-1H-indole-7-carboxylate

1-(Benzo[d][1,3]dioxol-5-yl)cyclopropanecarbonyl chloride (45 mg, 0.20mmol) and methyl 5-amino-2-tert-butyl-1H-indole-7-carboxylate (49.3 mg,0.200 mmol) were dissolved in N,N-dimethylformamide (2 mL) containing amagnetic stir bar and triethylamine (0.084 mL, 0.60 mmol). The resultingsolution was allowed to stir for 10 minutes at room temperature. Thecrude product was then purified by preparative HPLC using a gradient of0-99% acetonitrile in water containing 0.05% trifluoroacetic acid toyield methyl5-(1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarbox-amido)-2-tert-butyl-1H-indole-7-carboxylate.ESI-MS m/z calc. 434.2, found 435.5. (M+1)⁺. Retention time of 2.12minutes.

Example 67:1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-(1-hydroxy-2-methylpropan-2-yl)-1H-indol-5-yl)cyclopropanecarboxamide

To a solution of 1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxylic acid(0.075 g, 0.36 mmol) in acetonitrile (1.5 mL) were added HBTU (0.138 g,0.36 mmol) and Et₃N (152 μL, 1.09 mmol) at room temperature. The mixturewas stirred at room temperature for 10 minutes before a solution of2-(5-amino-1H-indol-2-yl)-2-methylpropan-1-ol (0.074 g, 0.36 mmol) inacetonitrile (1.94 mL) was added. After addition, the reaction mixturewas stirred at room temperature for 3 h. The solvent was evaporatedunder reduced pressure and the residue was dissolved in dichloromethane.The organic layer was washed with 1 N HCl (1×3 mL) and saturated aqueousNaHCO₃ (1×3 mL). The organic layer was dried over Na₂SO₄, filtered andevaporated under reduced pressure. The crude material was purified bycolumn chromatography on silica gel (ethyl acetate/hexane=1/1) to give1-(benzo[d][1,3]dioxol-5-yl)-N-(2-(1-hydroxy-2-methylpropan-2-yl)-1H-indol-5-yl)cyclopropanecarboxamide(0.11 g, 75%). ¹H NMR (400 MHz, DMSO-d6) δ 10.64 (s, 1H), 8.38 (s, 1H),7.55 (s, 1H), 7.15 (d, J=8.6 Hz, 1H), 7.04-6.90 (m, 4H), 6.06 (s, 1H),6.03 (s, 2H), 4.79 (t, J=2.7 Hz, 1H), 3.46 (d, J=0.0 Hz, 2H), 1.41-1.39(m, 2H), 1.26 (s, 6H), 1.05-1.02 (m, 2H).

Example 67:1-(Benzo[d][1,3]dioxol-5-yl)-N-(2,3,4,9-tetrahydro-1H-carbazol-6-yl)cyclopropanecarboxamide

2,3,4,9-Tetrahydro-1H-carbazol-6-amine (81.8 mg, 0.439 mmol) and1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxylic acid (90.4 mg, 0.439mmol) were dissolved in acetonitrile (3 mL) containingdiisopropylethylamine (0.230 mL, 1.32 mmol) and a magnetic stir bar.O-(7-Azabenzotriazol-1-yl)-N,N,N′,N′-tetramethyluroniumhexafluorophosphate (183 mg, 0.482 mmol) was added to the mixture andthe resulting solution was allowed to stir for 16 h at 70° C. Thesolvent was evaporated and the crude product was then purified on 40 gof silica gel utilizing a gradient of 5-50% ethyl acetate in hexanes toyield1-(benzo[d][1,3]dioxol-5-yl)-N-(2,3,4,9-tetrahydro-1H-carbazol-6-yl)cyclopropanecarboxamideas a beige powder (0.115 g, 70%) after drying. ESI-MS m z calc. 374.2,found 375.3 (M+1)⁺. Retention time of 3.43 minutes. ¹H NMR (400 MHz,DMSO-d6) δ 10.52 (s, 1H), 8.39 (s, 1H), 7.46 (d, J=1.8 Hz, 1H),7.10-6.89 (m, 5H), 6.03 (s, 2H), 2.68-2.65 (m, 2H), 2.56-2.54 (m, 2H),1.82-1.77 (m, 4H), 1.41-1.34 (m, 2H), 1.04-0.97 (m, 2H).

Example 69: tert-Butyl4-(5-(1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarbox-amido)-1H-indol-2-yl)piperidine-1-carboxylate

1-(Benzo[d][1,3]dioxol-5-yl)cyclopropanecarbonyl chloride (43 mg, 0.19mmol) and tert-butyl 4-(5-amino-1H-indol-2-yl)piperidine-1-carboxylate(60 mg, 0.19 mmol) were dissolved in dichloromethane (1 mL) containing amagnetic stir bar and triethylamine (0.056 mL, 0.40 mmol). The resultingsolution was allowed to stir for two days at room temperature. The crudeproduct was then evaporated to dryness, dissolved in a minimum ofN,N-dimethylformamide, and then purified by preparative HPLC using agradient of 0-99% acetonitrile in water containing 0.05% trifluoroaceticacid to yield tert-butyl4-(5-(1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1H-indol-2-yl)piperidine-1-carboxylate.ESI-MS m/z calc. 503.2, found 504.5. (M+1)⁺. Retention time of 1.99minutes.

Example 70: Ethyl2-(5-(1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1H-indol-2-yl)propanoate

tert-Butyl 2-(1-ethoxy-1-oxopropan-2-yl)-1H-indole-1-carboxylate

tert-Butyl 2-(2-ethoxy-2-oxoethyl)-1H-indole-1-carboxylate (3.0 g, 9.9mmol) was added to anhydrous THF (29 mL) and cooled to −78° C. A 0.5Msolution of potassium hexamethyldisilazane (20 mL, 9.9 mmol) was addedslowly such that the internal temperature stayed below −60° C. Stirringwas continued for 1 h at −78° C. Methyl iodide (727 μL, 11.7 mmol) wasadded to the mixture. The mixture was stirred for 30 minutes at roomtemperature. The mixture was quenched with sat. aq. ammonium chlorideand partitioned between water and dichloromethane. The aqueous phase wasextracted with dichloromethane and the combined organic phases weredried over Na₂SO₄ and evaporated under reduced pressure. The residue waspurified by column chromatography on silica gel(ethylacetate/hexane=1/9) to give tert-butyl2-(1-ethoxy-1-oxopropan-2-yl)-1H-indole-1-carboxylate (2.8 g, 88%).

Ethyl 2-(1H-indol-2-yl)propanoate

tert-Butyl 2-(1-ethoxy-1-oxopropan-2-yl)-1H-indole-1-carboxylate (2.77g, 8.74 mmol) was dissolved in dichloromethane (25 mL) before TFA (9.8mL) was added. The mixture was stirred for 1.5 h at room temperature.The mixture was evaporated to dryness, taken up in dichloromethane andwashed with sat. aq. sodium bicarbonate, water, and brine. The productwas purified by column chromatography on silica gel (0-20% EtOAc inhexane) to give ethyl 2-1H-indol-2-yl)propanoate (0.92 g, 50%).

Ethyl 2-(5-nitro-1H-indol-2-yl)propanoate

Ethyl 2-(1H-indol-2-yl)propanoate (0.91 g, 4.2 mmol) was dissolved inconcentrated sulfuric acid (3.9 mL) and cooled to −10° C.(salt/ice-mixture). A solution of sodium nitrate (0.36 g, 4.2 mmol) inconcentrated sulfuric acid (7.8 mL) was added dropwise over 35 min.Stirring was continued for another 30 min at −10° C. The mixture waspoured into ice and the product was extracted with ethyl acetate. Thecombined organic phases were washed with a small amount of sat. aq.sodium bicarbonate. The product was purified by column chromatography onsilica gel (5-30% EtOAc in hexane) to give ethyl2-(5-nitro-1H-indol-2-yl)propanoate (0.34 g, 31%).

Ethyl 2-(5-amino-1H-indol-2-yl)propanoate

To a solution of ethyl 2-(5-nitro-1H-indol-2-yl)propanoate (0.10 g, 0.38mmol) in ethanol (4 mL) was added tin chloride dihydrate (0.431 g, 1.91mmol). The mixture was heated in the microwave at 120° C. for 1 h. Themixture was diluted with ethyl acetate before water and saturatedaqueous NaHCO₃ were added. The reaction mixture was filtered through aplug of celite using ethyl acetate. The organic layer was separated fromthe aqueous layer. The organic layer was dried over Na₂SO₄, filtered andevaporated under reduced pressure to give ethyl2-(5-amino-1H-indol-2-yl)propanoate (0.088 g, 99%).

Ethyl2-(5-(1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1H-indol-2-yl)propanoate

To a solution of 1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxylic acid(0.079 g, 0.384 mmol) in acetonitrile (1.5 mL) were added HBTU (0.146 g,0.384 mmol) and Et₃N (160 μL, 1.15 mmol) at room temperature. Themixture was allowed to stir at room temperature for 10 min before asolution of ethyl 2-(5-amino-1H-indol-2-yl)propanoate (0.089 g, 0.384mmol) in acetonitrile (2.16 mL) was added. After addition, the reactionmixture was stirred at room temperature for 2 h. The solvent wasevaporated under reduced pressure and the residue was dissolved indichloromethane. The organic layer was washed with 1 N HCl (1×3 mL) andthen saturated aqueous NaHCO₃ (1×3 mL). The organic layer was dried overNa₂SO₄, filtered and evaporated under reduced pressure. The crudematerial was purified by column chromatography on silica gel (ethylacetate/hexane=1/1) to give ethyl 2-(5-(1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1H-indol-2-yl)propanoate(0.081 g, 50%). ¹H NMR (400 MHz, CDCl₃) δ 8.51 (s, 1H), 7.67 (s, 1H),7.23-7.19 (m, 2H), 7.04-7.01 (m, 3H), 6.89 (d, J=0.0 Hz, 1H), 6.28 (s,1H), 6.06 (s, 2H), 4.25-4.17 (m, 2H), 3.91 (q, J=7.2 Hz, 1H), 1.72-1.70(m, 2H), 1.61 (s, 2H), 1.29 (t, J=7.1 Hz, 4H), 1.13-1.11 (m, 2H).

Example 71: tert-Butyl2-(5-(1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarbox-amido)-1H-indol-2-yl)-2-methylpropylcarbamate

2-Methyl-2-(5-nitro-1H-indol-2-yl)propanoic Acid

Ethyl 2-methyl-2-(5-nitro-1H-indol-2-yl)propanoate (4.60 g, 16.7 mmol)was dissolved in THF/water (2:1, 30 mL). LiOH.H₂O (1.40 g, 33.3 mmol)was added and the mixture was stirred at 50° C. for 3 h. The mixture wasmade acidic by the careful addition of 3N HCl. The product was extractedwith ethylacetate and the combined organic phases were washed with brineand dried over magnesium sulfate to give2-methyl-2-(5-nitro-1H-indol-2-yl)propanoic acid (4.15 g, 99%).

2-Methyl-2-(5-nitro-1H-indol-2-yl)propanamide

2-Methyl-2-(5-nitro-1H-indol-2-yl)-propanoic acid (4.12 g, 16.6 mmol)was dissolved in acetonitrile (80 mL). EDC (3.80 g, 0.020 mmol), HOBt(2.70 g, 0.020 mmol), Et₃N (6.9 mL, 0.050 mmol) and ammonium chloride(1.34 g, 0.025 mmol) were added and the mixture was stirred overnight atroom temperature. Water was added and the mixture was extracted withethylacetate. Combined organic phases were washed with brine, dried overmagnesium sulfate and dried to give2-methyl-2-(5-nitro-1H-indol-2-yl)propanamide (4.3 g, 99%).

2-Methyl-2-(5-nitro-1H-indol-2-yl)propan-1-amine

2-Methyl-2-(5-nitro-1H-indol-2-yl)propanamide (200 mg, 0.81 mmol) wassuspended in THF (5 ml) and cooled to 0° C. Borane-THF complex solution(1.0 M, 2.4 mL, 2.4 mmol) was added slowly and the mixture was allowedto stir overnight at room temperature. The mixture was cooled to 0° C.and carefully acidified with 3 N HCl. THF was evaporated off, water wasadded and the mixture was washed with ethylacetate. The aqueous layerwas made alkaline with 50% NaOH and the mixture was extracted withethylacetate. The combined organic layers were dried over magnesiumsulfate, filtered and evaporated to give2-methyl-2-(5-nitro-1H-indol-2-yl)propan-1-amine (82 mg, 43%).

tert-Butyl 2-methyl-2-(5-nitro-1H-indol-2-yl)propylcarbamate

2-Methyl-2-(5-nitro-1H-indol-2-yl)propan-1-amine (137 mg, 0.587 mmol)was dissolved in THF (5 mL) and cooled to 0° C. Et₃N (82 μL, 0.59 mmol)and di-tert-butyl dicarbonate (129 mg, 0.587 mmol) were added and themixture was stirred at room temperature overnight. Water was added andthe mixture was extracted with ethylacetate. The residue was purified bysilica gel chromatography (10-40% ethylacetate in hexane) to givetert-butyl 2-methyl-2-(5-nitro-1H-indol-2-yl)propylcarbamate (131 mg,67%).

tert-Butyl 2-(5-amino-1H-indol-2-yl)-2-methylpropylcarbamate

To a solution of tert-butyl2-methyl-2-(5-nitro-1H-indol-2-yl)propylcarbamate (80 mg, 0.24 mmol) inTHF (9 mL) and water (2 mL) was added ammonium formate (60 mg, 0.96mmol) followed by 10% Pd/C (50 mg). The mixture was stirred at roomtemperature for 45 minutes. Pd/C was filtered off and the organicsolvent was removed by evaporation. The remaining aqueous phase wasextracted with dichloromethane. The combined organic phases were driedover magnesium sulfate and evaporated to give tert-butyl2-(5-amino-1H-indol-2-yl)-2-methylpropylcarbamate (58 mg, 80%).

tert-Butyl2-(5-(1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1H-indol-2-yl)-2-methylpropylcarbamate

tert-Butyl 2-(5-amino-1H-indol-2-yl)-2-methylpropylcarbamate (58 mg,0.19 mmol), 1-(benzo[d][1,3]dioxol-6-yl)cyclopropanecarboxylic acid (47mg, 0.23 mmol), EDC (45 mg, 0.23 mmol), HOBt (31 mg, 0.23 mmol) and Et₃N(80 μL, 0.57 mmol) were dissolved in DMF (4 mL) and stirred overnight atroom temperature. The mixture was diluted with water and extracted withethylacetate. The combined organic phases were dried over magnesiumsulfate and evaporated to dryness. The residue was purified by silicagel chromatography (10-30% ethylacetate in hexane) to give tert-butyl2-(5-(1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1H-indol-2-yl)-2-methylpropyl-carbamate(88 mg, 94%). ¹H NMR (400 MHz, CDCl₃) δ 8.32 (s, 1H), 7.62 (d, J=1.5 Hz,1H), 7.18-7.16 (m, 2H), 7.02-6.94 (m, 3H), 6.85 (d, J=7.8 Hz, 1H), 6.19(d, J=1.5 Hz, 1H), 6.02 (s, 2H), 4.54 (m, 1H), 3.33 (d, J=6.2 Hz, 2H),1.68 (dd, J=3.7, 6.8 Hz, 2H), 1.36 (s, 9H), 1.35 (s, 6H), 1.09 (dd,J=3.7, 6.8 Hz, 2H).

Example 72:(R)-N-(2-tert-Butyl-1-(2,3-dihydroxypropyl)-1H-indol-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide

(R)-2-tert-Butyl-1-((2,2-dimethyl-1,3-dioxolan-4-yl)methyl)-5-nitro-1H-indole

To a stirred solution of (S)-(2,2-dimethyl-1,3-dioxolan-4-yl)methyl4-methylbenzenesulfonate (1.58 g, 5.50 mmol) in anhydrous DMF (10 mL)under nitrogen gas was added 2-tert-butyl-5-nitro-1H-indole (1.00 g,4.58 mmol) followed by Cs₂CO₃ (2.99 g, 9.16 mol). The mixture wasstirred and heated at 80° C. under nitrogen gas. After 20 hours, 50%conversion was observed by LCMS. The reaction mixture was re-treatedwith Cs₂CO₃ (2.99 g, 9.16 mol) and(S)-(2,2-dimethyl-1,3-dioxolan-4-yl)methyl 4-methylbenzenesulfonate(1.58 g, 5.50 mmol) and heated at 80° C. for 24 hours. The reactionmixture was cooled to room temperature. The solids were filtered andwashed with ethyl acetate and hexane (1:1). The layers were separatedand the organic layer was washed with water (2×10 mL) and brine (2×10mL). The organic layer was dried over Na₂SO₄, filtered and evaporatedunder reduced pressure. The residue was purified by columnchromatography on silica gel (dichloromethane/hexane=1.5/1) to give(R)-2-tert-butyl-1-((2,2-dimethyl-1,3-dioxolan-4-yl)methyl)-5-nitro-1H-indole(1.0 g, 66%). ¹H NMR (400 MHz, CDCl₃) δ 8.48 (d, J=2.2 Hz, 1H), 8.08(dd, J=2.2, 9.1 Hz, 1H), 7.49 (d, J=9.1 Hz, 1H), 6.00 (s, 1H), 4.52-4.45(m, 3H), 4.12 (dd, J=6.0, 8.6 Hz, 1H), 3.78 (dd, J=6.0, 8.6 Hz, 1H),1.53 (s, 3H), 1.51 (s, 9H), 1.33 (s, 3H).

(R)-2-tert-Butyl-1-((2,2-dimethyl-1,3-dioxolan-4-yl)methyl-1H-indol-5-amine

To a stirred solution of(R)-2-tert-butyl-1-((2,2-dimethyl-1,3-dioxolan-4-yl)methyl)-5-nitro-1H-indole(1.0 g, 3.0 mmol) in ethanol (20 mL) and water (5 mL) was added ammoniumformate (0.76 g, 12 mmol) followed by slow addition of 10% palladium oncarbon (0.4 g). The mixture was stirred at room temperature for 1 h. Thereaction mixture was filtered through a plug of celite and rinsed withethyl acetate. The filtrate was evaporated under reduced pressure andthe crude product was dissolved in ethyl acetate. The organic layer waswashed with water (2×5 mL) and brine (2×5 mL). The organic layer wasdried over Na₂SO₄, filtered and evaporated under reduced pressure togive(R)-2-tert-butyl-1-((2,2-dimethyl-1,3-dioxolan-4-yl)methyl-1H-indol-5-amine(0.89 g, 98%). ¹H NMR (400 MHz, CDCl₃) δ 7.04 (d, J=4 Hz, 1H), 6.70 (d,J=2.2 Hz, 1H), 6.48 (dd, J=2.2, 8.6 Hz, 1H), 6.05 (s, 1H,), 4.38-4.1 (m,2H), 4.21 (dd, J=7.5, 16.5 Hz, 1H), 3.87 (dd, J=6.0, 8.6 Hz, 1H), 3.66(dd, J=6.0, 8.6 Hz, 1H), 3.33 (br s, 2H), 1.40 (s, 3H), 1.34 (s, 9H),1.25 (s, 3H).

N-((R)-2-tert-Butyl-1-((2,2-dimethyl-1,3-dioxolan-4-yl)methyl)-1H-indol-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide

To 1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxylic acid (0.73 g, 3.0mmol) was added thionyl chloride (660 μL, 9.0 mmol) and DMF (20 μL) atroom temperature. The mixture was stirred for 30 minutes before theexcess thionyl chloride was evaporated under reduced pressure. To theresulting acid chloride, dichloromethane (6.0 mL) and Et₃N (2.1 mL, 15mmol) were added. A solution of(R)-2-tert-butyl-1-((2,2-dimethyl-1,3-dioxolan-4-yl)methyl-1H-indol-5-amine(3.0 mmol) in dichloromethane (3.0 mL) was added to the cooled acidchloride solution. After addition, the reaction mixture was stirred atroom temperature for 45 minutes. The reaction mixture was filtered andthe filtrate was evaporated under reduced pressure. The residue waspurified by column chromatography on silica gel (ethylacetate/hexane=3/7) to giveN-((R)-2-tert-butyl-1-((2,2-dimethyl-1,3-dioxolan-4-yl)methyl)-1H-indol-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide(1.33 g, 84%). ¹H NMR (400 MHz, CDCl₃) δ 7.48 (d, J=2 Hz, 1H,), 7.31(dd, J=2, 8 Hz, 1H), 7.27 (dd, J=2, 8 Hz, 1H), 7.23 (d, J=8 Hz, 1H),7.14 (d, J=8 Hz, 1H), 7.02 (dd, J=2, 8 Hz, 1H), 6.92 (br s, 1H), 6.22(s, 1H), 4.38-4.05 (m, 3H), 3.91 (dd, J=5, 8 Hz, 1H), 3.75 (dd, J=5, 8Hz, 1H), 2.33 (q, J=8 Hz, 2H), 1.42 (s, 3H), 1.37 (s, 9H), 1.22 (s, 3H),1.10 (q, J=8 Hz, 2H).

N-((R)-2-tert-Butyl-1-((2,3-dihydroxypropyl)-1H-indol-5-yl)-1-(2,2-difluorobenzo-[d][1,3]dioxol-5-yl)cyclopropanecarboxamide

To a stirred solution ofN-(2-tert-butyl-1-((2,2-dimethyl-1,3-dioxolan-4-yl)methyl)-1H-indol-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide(1.28 g, 2.43 mmol) in methanol (34 mL) and water (3.7 mL) was addedpara-toluenesulfonic acid-hydrate (1.87 g, 9.83 mmol). The reactionmixture was stirred and heated at 80° C. for 25 minutes. The solvent wasevaporated under reduced pressure. The crude product was dissolved inethyl acetate. The organic layer was washed with saturated aqueousNaHCO₃ (2×10 mL) and brine (2×10 mL). The organic layer was dried overNa₂SO₄, filtered and evaporated under reduced pressure. The residue waspurified by column chromatography on silica gel (ethylacetate/hexane=13/7) to giveN-((R)-2-tert-butyl-1-((2,3-dihydroxypropyl)-1H-indol-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide(0.96 g, 81%). ¹H NMR (400 MHz, CDCl₃) δ 7.50 (d, J=2 Hz, 1H), 7.31 (dd,J=2, 8 Hz, 1H), 7.27 (dd, J=2, 8 Hz, 1H), 7.23 (d, J=8 Hz, 1H), 7.14 (d,J=8 Hz, 1H), 7.02 (br s, 1H,), 6.96 (dd, J=2, 8 Hz, 1H), 6.23 (s, 1H),4.35 (dd, J=8, 15 Hz, 1H), 4.26 (dd, J=4, 15 Hz, 1H,), 4.02-3.95 (m,1H), 3.60 (dd, J=4, 11 Hz, 1H), 3.50 (dd, J=5, 11 Hz, 1H), 1.75 (q, J=8Hz, 3H), 1.43 (s, 9H), 1.14 (q, J=8 Hz, 3H).

Example 73:3-(2-tert-Butyl-5-(1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1H-indol-1-yl)-2-hydroxypropanoicAcid

3-(2-tert-Butyl-5-(1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarbox-amido)-1H-indol-1-yl)-2-oxopropanoicAcid

To a solution ofN-(2-tert-butyl-1-(2,3-dihydroxypropyl)-1H-indol-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropane-carboxamide(97 mg, 0.20 mmol) in DMSO (1 mL) was added Dess-Martin periodinane (130mg, 0.30 mmol). The mixture was stirred at room temperature for 3 h. Thesolid was filtered off and washed with EtOAc. The filtrate waspartitioned between EtOAc and water. The aqueous layer was extractedwith EtOAc twice and the combined organic layers were washed with brineand dried over MgSO₄. After the removal of solvent, the residue waspurified by preparative TLC to yield3-(2-tert-butyl-5-(1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1H-indol-1-yl)-2-oxopropanoicacid that was used without further purification.

3-(2-tert-Butyl-5-(1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarbox-amido)-1H-indol-1-yl)-2-hydroxypropanoicAcid

To a solution of3-(2-tert-butyl-5-(1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1H-indol-1-yl)-2-oxopropanoicacid (50 mg, 0.10 mmol) in MeOH (1 mL) was added NaBH₄ (19 mg, 0.50mmol) at 0° C. The mixture was stirred at room temperature for 15 min.The resulting mixture was partitioned between EtOAc and water. Theaqueous layer was extracted with EtOAc twice and the combined organiclayers were washed with brine and dried over anhydrous MgSO₄. After theremoval of the solvent, the residue was taken up in DMSO and purified bypreparative LC/MS to give3-(2-tert-butyl-5-(1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1H-indol-1-yl)-2-hydroxypropanoicacid. ¹H NMR (400 MHz, CDCl₃) δ 7.36 (s), 7.27-7.23 (m, 2H), 7.15-7.11(m, 2H), 6.94 (d, J=8.5 Hz, 1H), 6.23 (s, 1H), 4.71 (s, 3H), 4.59 (q,J=10.3 Hz, 1H), 4.40-4.33 (m, 2H), 1.70 (d, J=1.9 Hz, 2H), 1.15 (q,J=4.0 Hz, 2H). ¹³C NMR (400 MHz, CDCl₃) δ 173.6, 173.1, 150.7, 144.1,143.6, 136.2, 135.4, 134.3, 131.7, 129.2, 129.0, 127.6, 126.7, 116.6,114.2, 112.4, 110.4, 110.1, 99.7, 70.3, 48.5, 32.6, 30.9, 30.7, 16.8. MS(ESI) m/e (M+H⁺) 501.2.

Example 74:(R)-N-(2-tert-Butyl-1-(2,3-dihydroxypropyl)-1H-indol-5-yl)-1-(2,2-dideuteriumbenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide

Methyl 1-(3,4-dihydroxyphenyl)cyclopropanecarboxylate

To a solution of 1-(3,4-dihydroxyphenyl)cyclopropanecarboxylic acid (190mg, 1.0 mmol) in MeOH (3 mL) was added 4-methylbenzenesulfonic acid (19mg, 0.10 mmol). The mixture was heated at 80° C. overnight. The reactionmixture was concentrated in vacuo and partitioned between EtOAc andwater. The aqueous layer was extracted with EtOAc twice and the combinedorganic layers were washed with sat. NaHCO₃ and brine and dried overMgSO₄. After the removal of solvent, the residue was dried in vacuo toyield methyl 1-(3,4-dihydroxyphenyl)cyclopropanecarboxylate (190 mg,91%) that was used without further purification. ¹H NMR (400 MHz,DMSO-d⁶) δ 6.76-6.71 (m, 2H), 6.66 (d, J=7.9 Hz, 1H), 3.56 (s, 3H), 1.50(q, J=3.6 Hz, 2H), 1.08 (q, J=3.6 Hz, 2H).

Methyl1-(2,2-dideuteriumbenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxylate

To a solution of methyl 1-(3,4-dihydroxyphenyl)cyclopropanecarboxylate(21 mg, 0.10 mmol) and CD₂Br₂ (35 mg, 0.20 mmol) in DMF (0.5 mL) wasadded Cs₂CO₃ (19 mg, 0.10 mmol). The mixture was heated at 120° C. for30 min. The reaction mixture was partitioned between EtOAc and water.The aqueous layer was extracted with EtOAc twice and the combinedorganic layers were washed with 1N NaOH and brine before being driedover MgSO₄. After the removal of solvent, the residue was dried in vacuoto yield methyl1-(2,2-dideuteriumbenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxylate (22mg) that was used without further purification. ¹H NMR (400 MHz, CDCl₃)δ 6.76-6.71 (m, 2H), 6.66 (d, J=7.9 Hz, 1H), 3.56 (s, 3H), 1.50 (q,J=3.6 Hz, 2H), 1.08 (q, J=3.6 Hz, 2H).

1-(2,2-Dideuteriumbenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxylic Acid

To a solution of methyl1-(2,2-dideuteriumbenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxylate (22mg, 0.10 mmol) in THF (0.5 mL) was added NaOH (1N, 0.25 mL, 0.25 mmol).The mixture was heated at 80° C. for 2 h. The reaction mixture waspartitioned between EtOAc and 1N NaOH. The aqueous layer was extractedwith EtOAc twice, neutralized with 1N HCl and extracted with EtOActwice. The combined organic layers were washed with brine and dried overMgSO₄. After the removal of solvent, the residue was dried in vacuo toyield 1-(2,2-dideuteriumbenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxylicacid (21 mg) that was used without further purification.

(R)-N-(2-tert-Butyl-1-((2,2-dimethyl-1,3-dioxolan-4-yl)methyl)-1H-indol-5-yl)-1-(2,2-dideuteriumbenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide

To a solution of1-(2,2-dideuteriumbenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxylic acid(21 mg, 0.10 mmol),(R)-2-tert-butyl-1-((2,2-dimethyl-1,3-dioxolan-4-yl)methyl)-1H-indol-5-amine(30 mg, 0.10 mmol), HATU (42 mg, 0.11 mol) in DMF (1 mL) was addedtriethylamine (0.030 mL, 0.22 mmol). The mixture was heated at roomtemperature for 5 min. The reaction mixture was partitioned betweenEtOAc and water. The aqueous layer was extracted with EtOAc twice andthe combined organic layers were washed with 1N NaOH, 1N HCl, and brinebefore being dried over MgSO₄. After the removal of solvent, the residuewas purified by column chromatography (20-40% ethyl acetate/hexane) toyield(R)-N-(2-tert-butyl-1-((2,2-dimethyl-1,3-dioxolan-4-yl)methyl)-1H-indol-5-yl)-1-(2,2-dideuteriumbenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide(24 mg, 49% from methyl 1-(3,4-dihydroxyphenyl)cyclopropanecarboxylate).MS (ESI) m/e (M+H⁺) 493.5.

(R)-N-(2-tert-Butyl-1-(2,3-dihydroxypropyl)-1H-indol-5-yl)-1-(2,2-dideuterium-benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide

To a solution of(R)-N-(2-tert-butyl-1-((2,2-dimethyl-1,3-dioxolan-4-yl)methyl)-1H-indol-5-yl)-1-(2,2-dideuterium-benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide(24 mg, 0.050 mmol), in methanol (0.5 mL) and water (0.05 mL) was added4-methylbenzenesulfonic acid (2.0 mg, 0.010 mmol). The mixture washeated at 80° C. for 30 min. The reaction mixture was partitionedbetween EtOAc and water. The aqueous layer was extracted with EtOActwice and the combined organic layers were washed with sat. NaHCO₃ andbrine before being dried over MgSO₄. After the removal of solvent, theresidue was purified by preparative HPLC to yield(R)-N-(2-tert-butyl-1-((2,2-dimethyl-1,3-dioxolan-4-yl)methyl)-1H-indol-5-yl)-1-(2,2-dideuteriumbenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide(12 mg, 52%). ¹H NMR (400 MHz, CDCl₃) δ 7.44 (d, J=2.0 Hz, 1H), 7.14(dd, J=22.8, 14.0 Hz, 2H), 6.95-6.89 (m, 2H), 6.78 (d, J=7.8 Hz, 1H),6.14 (s, 1H), 4.28 (dd, J=15.1, 8.3 Hz, 1H), 4.19 (dd, J=15.1, 4.5 Hz,1H), 4.05 (q, J=7.1 Hz, 1H), 3.55 (dd, J=11.3, 4.0 Hz, 1H), 3.45 (dd,J=11.3, 5.4 Hz, 1H), 1.60 (q, J=3.5 Hz, 2H), 1.35 (s, 9H), 1.02 (q,J=3.5 Hz, 2H). ¹³C NMR (400 MHz, CDCl₃) δ 171.4, 149.3, 147.1, 146.5,134.8, 132.3, 129.2, 126.5, 123.6, 114.3, 111.4, 110.4, 109.0, 107.8,98.5, 70.4, 63.1, 46.6, 31.6, 30.0, 29.8, 15.3. MS (ESI) m/e (M+H⁺)453.5.

It is further noted that the mono-deuterated analogue for this compoundcan be synthesized by substitution the reagent CHDBR₂ for CD₂BR₂ andfollowing the procedures described in example 74. Furthermore,deuterated analogues of the compounds as described herein such as offormula I can be produced using known synthetic methods as well as themethodology described herein. The deuterated analogues include both diand mono-deuterated analogues of the compounds of the present invention.The di and mono deuterated analogues of the compounds exhibit measurableactivity when tested using the assays described below.

Example 75:4-(5-(1-(Benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1H-indol-2-yl)-4-methylpentanoicAcid

1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-(4-cyano-2-methylbutan-2-yl)-1H-indol-5-yl)cyclopropanecarboxamide

To 1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxylic acid (0.068 g,0.33 mmol) was added thionyl chloride (72 μL, 0.99 mmol) and DMF (20 μL)at room temperature. The mixture was stirred for 30 minutes before theexcess thionyl chloride was evaporated under reduced pressure. To theresulting acid chloride, dichloromethane (0.5 mL) and Et₃N (230 μL, 1.7mmol) were added. A solution of4-(5-amino-1H-indol-2-yl)-4-methylpentanenitrile (0.33 mmol) indichloromethane (0.5 mL) was added to the acid chloride solution and themixture was stirred at room temperature for 1.5 h. The resulting mixturewas diluted with dichloromethane and washed with 1 N HCl (2×2 mL),saturated aqueous NaHCO₃ (2×2 mL) and brine (2×2 mL). The organic layerwas dried over anhydrous Na₂SO₄ and evaporated under reduced pressure togive1-(benzo[d][1,3]dioxol-5-yl)-N-(2-(4-cyano-2-methylbutan-2-yl)-1H-indol-5-yl)cyclopropanecarboxamide.

4-(5-(1-(Benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1H-indol-2-yl)-4-methylpentanoicAcid

A mixture of1-(benzo[d][1,3]dioxol-5-yl)-N-(2-(4-cyano-2-methylbutan-2-yl)-1H-indol-5-yl)cyclopropanecarboxamide(0.060 g, 0.15 mmol) and KOH (0.081 g, 1.5 mmol) in 50% EtOH/water (2mL) was heated in the microwave at 100° C. for 1 h. The solvent wasevaporated under reduced pressure. The crude product was dissolved inDMSO (1 mL), filtered, and purified by reverse phase preparative HPLC togive4-(5-(1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1H-indol-2-yl)-4-methylpentanoicacid. ¹H NMR (400 MHz, DMSO-d6) δ 11.98 (s, 1H), 10.79 (s, 1H), 8.44 (s,1H), 7.56 (s, 1H), 7.15 (d, J=8.6 Hz, 1H), 7.03-6.90 (m, 4H), 6.05 (s,1H), 6.02 (s, 2H), 1.97-1.87 (m, 4H), 1.41-1.38 (m, 2H), 1.30 (s, 6H),1.04-1.02 (m, 2H).

Example 76:1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-(1-hydroxypropan-2-yl)-1H-indol-5-yl)cyclopropanecarboxamide

2-(5-Nitro-1H-indol-2-yl)propan-1-ol

To a cooled solution of LiAlH₄ (1.0 M in THF, 1.2 mL, 1.2 mmol) in THF(5.3 mL) at 0° C. was added a solution of ethyl2-(5-nitro-1H-indol-2-yl)propanoate (0.20 g, 0.76 mmol) in THF (3.66 mL)dropwise. After addition, the mixture was allowed to warm up to roomtemperature and was stirred at room temperature for 3 h. The mixture wascooled to 0° C. Water (2 mL) was slowly added followed by carefuladdition of 15% NaOH (2 mL) and water (4 mL). The mixture was stirred atroom temperature for 0.5 h and was then filtered through a short plug ofcelite using ethyl acetate. The organic layer was separated from theaqueous layer, dried over Na₂SO₄, filtered and evaporated under reducedpressure. The residue was purified by column chromatography on silicagel (ethyl acetate/hexane=1/1) to give2-(5-nitro-1H-indol-2-yl)propan-1-ol (0.14 g, 81%).

2-(5-Amino-1H-indol-2-yl)propan-1-ol

To a solution of 2-(5-nitro-1H-indol-2-yl)propan-1-ol (0.13 g, 0.60mmol) in ethanol (5 mL) was added tin chloride dihydrate (0.67 g, 3.0mmol). The mixture was heated in the microwave at 120° C. for 1 h. Themixture was diluted with ethyl acetate before water and saturatedaqueous NaHCO₃ were added. The reaction mixture was filtered through aplug of celite using ethyl acetate. The organic layer was separated fromthe aqueous layer, dried over Na₂SO₄, filtered and evaporated underreduced pressure to give 2-(5-amino-1H-indol-2-yl)propan-1-ol (0.093 g,82%).

1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-(1-hydroxypropan-2-yl)-1H-indol-5-yl)cyclopropanecarboxamide

To a solution of 1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxylic acid(0.10 g, 0.49 mmol) in acetonitrile (2.0 mL) were added HBTU (0.185 g,0.49 mmol) and Et₃N (205 μL, 1.47 mmol) at room temperature. The mixturewas allowed to stir at room temperature for 10 minutes before a slurryof 2-(5-amino-1H-indol-2-yl)propan-1-ol (0.093 g, 0.49 mmol) inacetonitrile (2.7 mL) was added. After addition, the reaction mixturewas stirred at room temperature for 5.5 h. The solvent was evaporatedunder reduced pressure and the residue was dissolved in dichloromethane.The organic layer was washed with 1 N HCl (1×3 mL) and saturated aqueousNaHCO₃ (1×3 mL). The organic layer was dried over Na₂SO₄, filtered andevaporated under reduced pressure. The crude material was purified bycolumn chromatography on silica gel (ethyl acetate/hexane=13/7) to give1-(benzo[d][1,3]dioxol-5-yl)-N-(2-(1-hydroxypropan-2-yl)-1H-indol-5-yl)cyclopropanecarboxamide(0.095 g, 51%). H NMR (400 MHz, DMSO-d6) δ 10.74 (s, 1H), 8.38 (s, 1H),7.55 (s, 1H), 7.14 (d, J=8.6 Hz, 1H), 7.02-6.90 (m, 4H), 6.06 (s, 1H),6.02 (s, 2H), 4.76 (t, J=5.3 Hz, 1H), 3.68-3.63 (m, 1H), 3.50-3.44 (m,1H), 2.99-2.90 (m, 1H), 1.41-1.38 (m, 2H), 1.26 (d, J=7.0 Hz, 3H),1.05-1.02 (m, 2H).

Example 77:1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-1H-indol-5-yl)-N-methylcyclopropanecarboxamide

1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-1H-indol-5-yl)-N-methylcyclopropanecarboxamide

2-tert-Butyl-N-methyl-1H-indol-5-amine (20.2 mg, 0.100 mmol) and1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxylic acid (20.6 mg, 0.100mmol) were dissolved in N,N-dimethylformamide (1 mL) containingtriethylamine (42.1 μL, 0.300 mmol) and a magnetic stir bar.O-(7-Azabenzotriazol-1-yl)-N,N,N′,N′-tetramethyluroniumhexafluorophosphate (42 mg, 0.11 mmol) was added to the mixture and theresulting solution was allowed to stir for 16 h at 80° C. The crudeproduct was then purified by preparative HPLC utilizing a gradient of0-99% acetonitrile in water containing 0.05% trifluoroacetic acid toyield1-(benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-1H-indol-5-yl)-N-methylcyclopropanecarboxamide.ESI-MS m/z calc. 390.2, found 391.3 (M+1)⁺. Retention time of 3.41minutes.

Example 78:N-(2-tert-Butyl-1-methyl-1H-indol-5-yl)-1-(benzo[d][1,3]dioxol-6-yl)-N-methylcyclopropanecarboxamide

Sodium hydride (0.028 g, 0.70 mmol, 60% by weight dispersion in oil) wasslowly added to a stirred solution ofN-(2-tert-butyl-1H-indol-5-yl)-1-(benzo[d][1,3]dioxol-6-yl)cyclopropanecarboxamide(0.250 g, 0.664 mmol) in a mixture of 4.5 mL of anhydroustetrahydrofuran (THF) and 0.5 mL of anhydrous N,N-dimethylformamide(DMF). The resulting suspension was allowed to stir for 2 minutes andthen iodomethane (0.062 mL, 1.0 mmol) was added to the reaction mixture.Two additional aliquots of sodium hydride and iodomethane were requiredto consume all of the starting material which was monitored by LC/MS.The crude reaction product was evaporated to dryness, redissolved in aminimum of DMF and purified by preparative LC/MS chromatography to yieldthe pure product (0.0343 g, 13%) ESI-MS m/z calc. 404.2, found 405.3(M+1)⁺. Retention time of 3.65 minutes.

Example 79:1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-(hydroxymethyl)-1H-indol-5-yl)cyclopropanecarboxamide

Ethyl5-(1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1H-indole-2-carboxylate(1.18 g, 3.0 mmol) was added to a solution of LiBH₄ (132 mg, 6.0 mmol)in THF (10 mL) and water (0.1 mL). The mixture was allowed to stir for16 h at 25° C. before it was quenched with water (10 mL) and slowly madeacidic by addition of 1 N HCl. The mixture was extracted with three50-mL portions of ethyl acetate. The organic extracts were dried overNa₂SO₄ and evaporated to yield1-(benzo[d][1,3]dioxol-5-yl)-N-(2-(hydroxymethyl)-1H-indol-5-yl)cyclopropanecarboxamide(770 mg, 73%). A small amount was further purified by reverse phaseHPLC. ESI-MS m/z calc. 350.4, found 351.3 (M+1)⁺; retention time 2.59minutes.

Example 80:5-(1-(Benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-N-tert-butyl-1H-indole-2-carboxamide

5-(1-(Benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1H-indole-2-carboxylicAcid

Ethyl5-(1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1H-indole-2-carboxylate(392 mg, 1.0 mmol) and LiOH (126 mg, 3 mmol) were dissolved in H₂O (5mL) and 1,4-dioxane (3 mL). The mixture was heated in an oil bath at100° C. for 24 hours before it was cooled to room temperature. Themixture was acidified with 1N HCl and it was extracted with three 20 mLportions of dichloromethane. The organic extracts were dried over Na₂SO₄and evaporated to yield5-(1-(benzo[d][1,3]-dioxol-5-yl)cyclopropanecarboxamido)-1H-indole-2-carboxylicacid (302 mg, 83%). A small amount was further purified by reverse phaseHPLC. ESI-MS m/z calc. 364.1, found 365.1 (M+1)⁺; retention time 2.70minutes.

5-(1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-N-tert-butyl-1H-indole-2-carboxamide

5-(1-(Benzo[d][1,3]dioxol-5-yl)cyclopropane-carboxamido)-1H-indole-2-carboxylicacid (36 mg, 0.10 mmol) and 2-methylpropan-2-amine (8.8 mg, 0.12 mmol)were dissolved in N,N-dimethylformamide (1.0 mL) containingtriethylamine (28 μL, 0.20 mmol).O-(7-Azabenzotriazol-1-yl)-N,N,N′,N′-tetramethyluroniumhexafluorophosphate (46 mg, 0.12 mmol) was added to the mixture and theresulting solution was allowed to stir for 3 hours. The mixture wasfiltered and purified by reverse phase HPLC to yield5-(1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-N-tert-butyl-1H-indole-2-carboxamide.ESI-MS m/z calc. 419.2, found 420.3 (M+1)⁺; retention time 3.12 minutes.

Example 81:N-(3-Amino-2-tert-butyl-1H-indol-5-yl)-1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide

A solution of1-(benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-1H-indol-5-yl)cyclopropanecarboxamide (50 mg, 0.13 mmol) was dissolved in AcOH (2 mL) and warmedto 45° C. To the mixture was added a solution of NaNO₂ (9 mg) in H₂O(0.03 mL). The mixture was allowed to stir for 30 min at 45° C. beforethe precipitate was collected and washed with Et₂O. This material wasused in the next step without further purification. To the crudematerial,1-(benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-3-nitroso-1H-indol-5-yl)cyclopropanecarboxamide,was added AcOH (2 mL) and Zn dust (5 mg). The mixture was allowed tostir for 1 h at ambient temperature. EtOAc and H₂O were added to themixture. The layers were separated and the organic layer was washed withsat. aq. NaHCO₃, dried over MgSO₄, and concentrated in vacuo. Theresidue was taken up in DMF (1 mL) and was purified using prep-HPLC.LCMS: m/z 392.3; retention time of 2.18 min.

Example 82:1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-3-(methylsulfonyl)-1H-indol-5-yl)cyclopropanecarboxamide

1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-3-(methylsulfonyl)-1H-indol-5-yl)cyclopropanecarboxamide

To a solution of1-(benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-1H-indol-5-yl)cyclopropanecarboxamide(120 mg, 0.31 mmol) in anhydrous DMF-THF (3.3 mL, 1:9) was added NaH(60% in mineral oil, 49 mg, 1.2 mmol) at room temperature. After 30 minunder N₂, the suspension was cooled down to −15° C. and a solution ofmethanesulfonyl chloride (1.1 eq.) in DMF (0.5 mL) was added dropwise.The reaction mixture was stirred for 30 min at −15° C. then for 6 h atroom temperature. Water (0.5 mL) was added at 0° C., solvent wasremoved, and the residue was diluted with MeOH, filtrated and purifiedby preparative HPLC to give1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-3-(methylsulfonyl)-1H-indol-5-yl)cyclopropanecarboxamide.¹H NMR (400 MHz, DMSO) δ 11.6 (s, 1H), 8.7 (s, 1H), 7.94 (d, J=1.7 Hz,1H), 7.38 (d, J=8.7 Hz, 1H), 7.33 (dd, J1=1.9 Hz, J2=8.7 Hz, 1H), 7.03(d, J=1.7 Hz, 1H), 6.95 (dd, J1=1.7 Hz, J2=8.0 Hz, 1H), 6.90 (d, J=8.0Hz, 1H), 6.02 (s, 2H), 3.07 (s, 3H), 1.56-1.40 (m, 9H), 1.41 (dd, J1=4.0Hz, J2=6.7 Hz, 2H), 1.03 (dd, J1=4.0 Hz, J2=6.7 Hz, 2H). MS (ESI) m/e(M+H⁺) 455.5.

Example 83:1-(Benzo[d][1,3]dioxol-5-yl)-N-(3-phenyl-1H-indol-5-yl)cyclopropaneCarboxamide

1-(Benzo[d][1,3]dioxol-5-yl)-N-(3-bromo-1H-indol-5-yl)cyclopropanecarboxamide

Freshly recrystallized N-bromosuccinimide (0.278 g, 1.56 mmol) was addedportionwise to a solution of1-(benzo[d][1,3]dioxol-5-yl)-N-(1H-indol-5-yl)cyclopropanecarboxamide(0.500 g, 1.56 mmol) in N,N-dimethylformamide (2 mL) over 2 minutes. Thereaction mixture was protected from light and was stirred bar for 5minutes. The resulting green solution was poured into 40 mL of water.The grey precipitate which formed was filtered and washed with water toyield1-(benzo[d][1,3]dioxol-5-yl)-N-(3-bromo-1H-indol-5-yl)cyclopropanecarboxamide(0.564 g, 91%). ESI-MS m/z calc. 398.0, found 399.3 (M+1)⁺. Retentiontime of 3.38 minutes. ¹H NMR (400 MHz, DMSO-d6) 11.37 (s, 1H), 8.71 (s,1H), 7.67 (d, J=1.8 Hz, 1H), 7.50 (d, J=2.6 Hz, 1H), 7.29 (d, J=8.8 Hz,1H), 7.22 (dd, J=2.0, 8.8 Hz, 1H), 7.02 (d, J=1.6 Hz, 1H), 6.96-6.88 (m,2H), 6.03 (s, 2H), 1.43-1.40 (m, 2H), 1.09-1.04 (m, 2H).

1-(Benzo[d][1,3]dioxol-5-yl)-N-(3-phenyl-1H-indol-5-yl)cyclopropanecarboxamide

Phenyl boronic acid (24.6 mg, 0.204 mmol) was added to a solution of1-(benzo[d][1,3]-dioxol-5-yl)-N-(3-bromo-1H-indol-5-yl)cyclopropanecarboxamide(39.9 mg, 0.100 mmol) in ethanol (1 mL) containing FibreCat 1001 (6 mg)and 1M aqueous potassium carbonate (0.260 mL). The reaction mixture wasthen heated at 130° C. in a microwave reactor for 20 minutes. The crudeproduct was then purified by preparative HPLC utilizing a gradient of0-99% acetonitrile in water containing 0.05% trifluoroacetic acid toyield1-(benzo[d][1,3]dioxol-5-yl)-N-(3-phenyl-1H-indol-5-yl)cyclopropanecarboxamide. ESI-MS m/z calc. 396.2, found 397.3 (M+1)⁺. Retention timeof 3.52 minutes. ¹H NMR (400 MHz, DMSO-d6) δ 11.27 (d, J=1.9 Hz, 1H),8.66 (s, 1H), 8.08 (d, J=1.6 Hz, 1H), 7.65-7.61 (m, 3H), 7.46-7.40 (m,2H), 7.31 (d, J=8.7 Hz, 1H), 7.25-7.17 (m, 2H), 7.03 (d, J=1.6 Hz, 1H),6.98-6.87 (m, 2H), 6.02 (s, 2H), 1.43-1.39 (m, 2H), 1.06-1.02 (m, 2H).

Example 84:1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-3-cyano-1H-indol-5-yl)cyclopropanecarboxamide

1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-3-formyl-1H-indol-5-yl)cyclopropane-carboxamide

POCl₃ (12 g, 80 mmol) was added dropwise to DMF (40 mL) held at −20° C.After the addition was complete, the reaction mixture was allowed towarm to 0° C. and was stirred for 1 h.1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-1H-indol-5-yl)cyclopropanecarboxamide(3.0 g, 8.0 mmol) was added and the mixture was warmed to 25° C. Afterstirring for 30 minutes the reaction mixture was poured over ice andstirred for 2 h. The mixture was then heated at 100° C. for 30 min. Themixture was cooled and the solid precipitate was collected and washedwith water. The solid was then dissolved in 200 mL dichloromethane andwashed with 200 mL of a saturated aq. NaHCO₃. The organics were driedover Na₂SO₄ and evaporated to yield1-(benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-3-formyl-1H-indol-5-yl)cyclopropane-carboxamide(2.0 g, 61%). ESI-MS m/z calc. 404.5, found 405.5 (M+1)⁺; retention time3.30 minutes. ¹H NMR (400 MHz, DMSO-d6) δ 11.48 (s, 1H), 10.39 (s, 1H),8.72 (s, 1H), 8.21 (s, 1H), 7.35-7.31 (m, 2H), 7.04-7.03 (m, 1H),6.97-6.90 (m, 2H), 6.03 (s, 2H), 1.53 (s, 9H), 1.42-1.39 (m, 2H),1.05-1.03 (m, 2H).

(Z)-1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-3-((hydroxyimino)methyl)-1H-indol-5-yl)cyclopropanecarboxamide

To a solution of1-(benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-3-formyl-1H-indol-5-yl)cyclopropanecarboxamide(100 mg, 0.25 mmol) in dichloromethane (5 mL) was added hydroxylaminehydrochloride (21 mg, 0.30 mmol). After stirring for 48 h, the mixturewas evaporated to dryness and purified by column chromatography (0-100%ethyl acetate/hexanes) to yield(Z)-1-(benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-3-((hydroxyimino)methyl)-1H-indol-5-yl)cyclopropanecarboxamide(81 mg, 77%). ESI-MS m/z calc. 419.5, found 420.5 (M+1)⁺; retention time3.42 minutes. ¹H NMR (400 MHz, DMSO-d6) δ 10.86 (s, 0.5H), 10.55 (s,0.5H), 8.56-8.50 (m, 2H), 8.02 (m, 1H), 7.24-7.22 (m, 1H), 7.12-7.10 (m,1H), 7.03 (m, 1H), 6.96-6.90 (m, 2H), 6.03 (s, 2H), 1.43 (s, 9H),1.40-1.38 (m, 2H), 1.04-1.01 (m, 2H).

1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-3-cyano-1H-indol-5-yl)cyclopropane-carboxamide

(Z)-1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-3-((hydroxyimino)-methyl)-1H-indol-5-yl)cyclopropanecarboxamide(39 mg, 0.090 mmol) was dissolved in acetic anhydride (1 mL) and heatedat reflux for 3 h. The mixture was cooled in an ice bath and theprecipitate was collected and washed with water. The solid was furtherdried under high vacuum to yield1-(benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-3-cyano-1H-indol-5-yl)cyclopropanecarboxamide.ESI-MS m/z calc. 401.5, found 402.5 (M+1)⁺; retention time 3.70 minutes.¹H NMR (400 MHz, DMSO-d6) δ 11.72 (s, 1H), 8.79 (s, 1H), 7.79 (s, 1H),7.32 (m, 2H), 7.03-7.02 (m, 1H), 6.95-6.89 (m, 2H), 6.03 (s, 2H), 1.47(s, 9H), 1.43-1.41 (m, 2H), 1.06-1.04 (m, 2H).

Example 85:1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-3-methyl-1H-indol-5-yl)cyclopropanecarboxamide

A solution of1-(benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-1H-indol-5-yl)cyclopropanecarboxamide(75 mg, 0.20 mmol) and iodomethane (125 μL, 2.0 mmol) inN,N-dimethylformamide (1 mL) was heated at 120° C. in a sealed tube for24 h. The reaction was filtered and purified by reverse phase HPLC.ESI-MS m/z calc. 390.5, found 391.3 (M+1)⁺; retention time 2.04 minutes.¹H NMR (400 MHz, DMSO-d6) δ 10.30 (s, 1H), 8.39 (s, 1H), 7.51 (m, 1H),7.13-7.11 (m, 1H), 7.03-6.90 (m, 4H), 6.03 (s, 2H), 2.25 (s, 3H),1.40-1.38 (m, 11H), 1.03-1.01 (m, 2H).

Example 86:1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-3-(2-hydroxyethyl)-1H-indol-5-yl)cyclopropanecarboxamide

Approximately 100 μL of ethylene dioxide was condensed in a reactiontube at −78° C. A solution of1-(benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-1H-indol-5-yl)cyclopropanecarboxamide(200 mg, 0.50 mmol) and indium trichloride (20 mg, 0.10 mmol) indichloromethane (2 mL) was added and the reaction mixture was irradiatedin the microwave for 20 min at 100° C. The volatiles were removed andthe residue was purified by column chromatography (0-100% ethylacetate/hexanes) to give1-(benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-3-(2-hydroxyethyl)-1H-indol-5-yl)cyclopropanecarboxamide(5 mg, 3%). ESI-MS m/z calc. 420.5, found 421.3 (M+1)⁺; retention time1.67 minutes. ¹H NMR (400 MHz, CD₃CN) δ 8.78 (s, 1H), 7.40 (m, 1H), 7.33(s, 1H), 7.08 (m, 1H), 6.95-6.87 (m, 3H), 6.79 (m, 1H), 5.91 (s, 2H),3.51 (dd, J=5.9, 7.8 Hz, 2H), 2.92-2.88 (m, 2H), 2.64 (t, J=5.8 Hz, 1H),1.50 (m, 2H), 1.41 (s, 9H), 1.06 (m, 2H).

Example 87:2-(5-(1-(Benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1H-indol-2-yl)aceticacid

To a solution of ethyl2-(5-(1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1H-indol-2-yl)acetate(0.010 g, 0.025 mmol) in THF (0.3 mL) were added LiOH.H₂O (0.002 g, 0.05mmol) and water (0.15 mL) were added. The mixture was stirred at roomtemperature for 2 h. dichloromethane (3 mL) was added to the reactionmixture and the organic layer was washed with 1 N HCl (2×1.5 mL) andwater (2×1.5 mL). The organic layer was dried over Na₂SO₄ and filtered.The filtrate was evaporated under reduced pressure to give2-(5-(1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1H-indol-2-yl)-aceticacid. ¹H NMR (400 MHz, DMSO-d6) δ 12.53 (s, 1H), 10.90 (s, 1H), 8.42 (s,1H), 7.57 (s, 1H), 7.17 (d, J=8.6 Hz, 1H), 7.05-6.90 (m, 4H), 6.17 (s,1H), 6.02 (s, 2H), 3.69 (s, 2H), 1.41-1.39 (m, 2H), 1.04-1.02 (m, 2H).

Example 88:5-(1-(Benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-2-tert-butyl-1H-indole-7-carboxylicAcid

Methyl5-(1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-2-tert-butyl-1H-indole-7-carboxylate(30 mg, 0.069 mmol) was dissolved in a mixture of 1,4-dioxane (1.5 mL)and water (2 mL) containing a magnetic star bar and lithium hydroxide(30 mg, 0.71 mmol). The resulting solution was stirred at 70° C. for 45minutes. The crude product was then acidified with 2.6 M hydrochloricacid and extracted three times with an equivalent volume ofdichloromethane. The dichloromethane extracts were combined, dried oversodium sulfate, filtered, and evaporated to dryness. The residue wasdissolved in a minimum of N,N-dimethylformamide and then purified bypreparative HPLC using a gradient of 0-99% acetonitrile in watercontaining 0.05% trifluoroacetic acid to yield5-(1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-2-tert-butyl-1H-indole-7-carboxylicacid. ESI-MS m/z calc. 434.2, found 435.5. Retention time of 1.85minutes. ¹H NMR (400 MHz, DMSO-d6) δ 13.05 (s, 1H), 9.96 (d, J=1.6 Hz,1H), 7.89 (d, J=1.9 Hz, 1H), 7.74 (d, J=2.0 Hz, 1H), 7.02 (d, J=1.6 Hz,1H), 6.96-6.88 (m, 2H), 6.22 (d, J=2.3 Hz, 1H), 6.02 (s, 2H), 1.43-1.40(m, 2H), 1.37 (s, 9H), 1.06-1.02 (m, 2H).

Example 89:1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-1-(1,3-dihydroxypropan-2-yl)-1H-indol-5-yl)cyclopropanecarboxamide

1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-1-(1,3-dihydroxypropan-2-yl)indolin-5-yl)cyclopropanecarboxamide

1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butylindolin-5-yl)cyclopropanecarboxamide(50 mg, 0.13 mmol) was dissolved in dichloroethane (0.20 mL) and2,2-dimethyl-1,3-dioxan-5-one (0.20 mL). Trifluoroacetic acid was added(0.039 mL) and the resulting solution was allowed to stir for 20minutes. Sodium triacetoxyborohydride was added (55 mg, 0.26 mmol) andthe reaction mixture was stirred for 30 minutes. The crude reactionmixture was then evaporated to dryness, dissolved inN,N-dimethylformamide and purified by preparative HPLC using a gradientof 0-99% acetonitrile in water containing 0.05% trifluoroacetic acid.

1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-1-(1,3-dihydroxypropan-2-yl)-1H-indol-5-yl)cyclopropanecarboxamide

1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-1-(1,3-dihydroxypropan-2-yl)indolin-5-yl)cyclopropanecarboxamide(40.3 mg, 0.0711 mmol as the trifluoracetic acid salt) was dissolved intoluene (1 mL). To the resulting solution was added2,3,5,6-tetrachlorocyclohexa-2,5-diene-1,4-dione (35 mg, 0.14 mmol). Theresulting suspension was heated at 100° C. in an oil bath for 10minutes. The crude product was then evaporated to dryness, dissolved ina 1 mL of N,N-dimethylformamide and purified by purified by preparativeHPLC using a gradient of 0-99% acetonitrile in water containing 0.05%trifluoroacetic acid to yield1-(benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-1-(1,3-dihydroxypropan-2-yl)-1H-indol-5-yl)cyclopropanecarboxamide.ESI-MS m/z calc. 450.2, found 451.5 (M+1)⁺. Retention time of 1.59minutes.

Example 90:N-(7-(Aminomethyl)-2-tert-butyl-1H-indol-5-yl)-1-(benzo[d][1,3]-dioxol-5-yl)cyclopropanecarboxamide

N-(7-(Aminomethyl)-2-tert-butyl-1H-indol-5-yl)-1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide

1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-7-cyano-1H-indol-5-yl)cyclopropanecarboxamide(375 mg, 0.934 mmol) was dissolved in 35 mL of ethyl acetate. Thesolution was recirculated through a continuous flow hydrogenationreactor containing 10% palladium on carbon at 100° C. under 100 bar ofhydrogen for 8 h. The crude product was then evaporated to dryness andpurified on 12 g of silica gel utilizing a gradient of 0-100% ethylacetate (containing 0.5% triethylamine) in hexanes to yieldN-(7-(aminomethyl)-2-tert-butyl-1H-indol-5-yl)-1-(benzo[d][1,3]-dioxol-5-yl)-cyclopropanecarboxamide(121 mg, 32%). ESI-MS m/z calc. 405.2, found 406.5 (M+1)⁺. Retentiontime of 1.48 minutes.

Example 91:5-(1-(Benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-2-tert-butyl-1H-indole-7-carboxamide

5-(1-(Benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-2-tert-butyl-1H-indole-7-carboxamide

1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-7-cyano-1H-indol-5-yl)-cyclopropanecarboxamide(45 mg, 0.11 mmol) was suspended in a mixture of methanol (1.8 mL), 30%aqueous hydrogen peroxide (0.14 mL, 4.4 mmol) and 10% aqueous sodiumhydroxide (0.150 mL). The resulting suspension was stirred for 72 h atroom temperature. The hydrogen peroxide was then quenched with sodiumsulfite. The reaction mixture was diluted with 0.5 mL ofN,N-dimethylformamide, filtered, and purified by preparative HPLC usinga gradient of 0-99% acetonitrile in water containing 0.05%trifluoroacetic acid to yield5-(1-(benzo[d][1,3]dioxol-5-yl)cyclopropane-carboxamido)-2-tert-butyl-1H-indole-7-carboxamide.ESI-MS m/z calc. 419.2, found 420.3 (M+1)⁺. Retention time of 1.74minutes.

Example 92:1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-7-(methylsulfonamido-methyl)-1H-indol-5-yl)cyclopropanecarboxamide

1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-7-(methylsulfonamidomethyl)-1H-indol-5-yl)cyclopropanecarboxamide

N-(7-(Aminomethyl)-2-tert-butyl-1H-indol-5-yl)-1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide(20 mg, 0.049 mmol) was dissolved in DMF (0.5 mL) containingtriethylamine (20.6 μL, 0.147 mmol) and a magnetic stir bar.Methanesulfonyl chloride (4.2 μL, 0.054 mmol) was then added to thereaction mixture. The reaction mixture was allowed to stir for 12 h atroom temperature. The crude product was purified by preparative HPLCusing a gradient of 0-99% acetonitrile in water containing 0.05%trifluoroacetic acid to yield1-(benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-7-(methylsulfonamidomethyl)-1H-indol-5-yl)cyclopropanecarboxamide.ESI-MS m/z calc. 483.2, found 484.3 (M+1)⁺. Retention time of 1.84minutes.

Example 93:N-(7-(Acetamidomethyl)-2-tert-butyl-1H-indol-5-yl)-1-(benzo[d][1,3]-dioxol-5-yl)cyclopropanecarboxamide

N-(7-(Aminomethyl)-2-tert-butyl-1H-indol-5-yl)-1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide(20 mg, 0.049 mmol) was dissolved in DMF (0.5 mL) containingtriethylamine (20.6 μL, 0.147 mmol) and a magnetic stir bar. Acetylchloride (4.2 μL, 0.054 mmol) was then added to the reaction mixture.The reaction mixture was allowed to stir for 16 h at room temperature.The crude product was purified by preparative HPLC using a gradient of0-99% acetonitrile in water containing 0.05% trifluoroacetic acid toyieldN-(7-(acetamidomethyl)-2-tert-butyl-1H-indol-5-yl)-1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide.ESI-MS m/z calc. 447.2, found 448.3 (M+1)⁺. Retention time of 1.76minutes.

Example 94:N-(1-Acetyl-2-tert-butyl-1H-indol-5-yl)-1-(benzo[d][1,3]dioxol-5-yl)-cyclopropanecarboxamide

To a solution of1-(benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-1H-indol-5-yl)cyclopropanecarboxamide(120 mg, 0.31 mmol) in anhydrous DMF-THF (3.3 mL, 1:9) was added NaH(60% in mineral oil, 49 mg, 1.2 mmol) at room temperature. After 30 minunder N₂, the suspension was cooled down to −15° C. and a solution ofacetyl chloride (1.1 eq.) in DMF (0.5 mL) was added dropwise. Thereaction mixture was stirred for 30 min at −15° C. then for 6 h at roomtemperature. Water (0.5 mL) was added at 0° C., solvent was removed, andthe residue was diluted with MeOH, filtrated and purified by preparativeHPLC to giveN-(1-acetyl-2-tert-butyl-1H-indol-5-yl)-1-(benzo[d][1,3]dioxol-5-yl)cyclo-propanecarboxamide.¹H NMR (400 MHz, DMSO) δ 8.9 (s, 1H), 7.74 (d, J=2.1 Hz, 1H), 7.54 (d,J=9.0 Hz, 1H), 7.28 (dd, J1=2.1 Hz, J2=9.0 Hz, 1H), 7.01 (d, J=1.5 Hz,1H), 6.93 (dd, J1=1.7 Hz, J2=8.0 Hz, 1H), 6.89 (d, J=8.0 Hz, 1H), 6.54(bs, 1H), 6.02 (s, 2H), 2.80 (s, 3H), 1.42-1.40 (m, 11H), 1.06-1.05 (m,2H). MS (ESI) m/e (M+H⁺) 419.3.

Example 95:N-(1-(2-Acetamidoethyl)-2-tert-butyl-6-fluoro-1H-indol-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide

N-(1-(2-Aminoethyl)-2-tert-butyl-6-fluoro-1H-indol-5-yl)-1-(2,2-difluorobenzo-[d][1,3]dioxol-5-yl)cyclopropanecarboxamide

To a solution of tert-butyl2-(2-tert-butyl-5-(1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-6-fluoro-1H-indol-1-yl)ethylcarbamate(620 mg, 1.08 mmol) in CH₂Cl₂ (8 mL) was added TFA (2 mL). The reactionwas stirred at room temperature for 1.5 h before being neutralized withsolid NaHCO₃. The solution was partitioned between H₂O and CH₂Cl₂. Theorganic layer was dried over MgSO₄, filtered and concentrated to yieldthe product as a cream colored solid (365 mg, 71%). ¹H NMR (400 MHz,DMSO-d6) δ 8.38 (s, 1H), 7.87 (br s, 3H, NH₃+), 7.52 (s, 1H), 7.45-7.38(m, 3H), 7.32 (dd, J=8.3, 1.5 Hz, 1H), 6.21 (s, 1H), 4.46 (m, 2H), 3.02(m, 2H), 1.46 (m, 2H), 1.41 (s, 9H), 1.14 (m, 2H). HPLC ret. time 1.66min, 10-99% CH₃CN, 3 min run; ESI-MS 474.4 m/z (M+H⁺).

N-(1-(2-Acetamidoethyl)-2-tert-butyl-6-fluoro-1H-indol-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide

To a solution ofN-(1-(2-aminoethyl)-2-tert-butyl-6-fluoro-1H-indol-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropane-carboxamide(47 mg, 0.10 mmol) and Et₃N (28 μL, 0.20 mmol) in DMF (1 mL) was addedacetyl chloride (7.1 μL, 0.10 mmol). The mixture was stirred at roomtemperature for 1 h before being filtered and purified by reverse phaseHPLC (10-99% CH₃CN/H₂O) to yieldN-(1-(2-acetamidoethyl)-2-tert-butyl-6-fluoro-1H-indol-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide.¹H NMR (400 MHz, DMSO-d6) δ 8.35 (s, 1H), 8.15 (t, J=5.9 Hz, 1H), 7.53(s, 1H), 7.43-7.31 (m, 4H), 6.17 (s, 1H), 4.22 (m, 2H), 3.30 (m, 2H),1.85 (s, 3H), 1.47 (m, 2H), 1.41 (s, 9H), 1.13 (m, 2H). HPLC ret. time2.06 min, 10-99% CH₃CN, 3 min run; ESI-MS 516.4 m/z (M+H⁺).

Example 96:1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-1-(2-hydroxy-3-methoxy-propyl)-1H-indol-5-yl)cyclopropenecarboxamide

1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-1H-indol-5-yl)cyclopropanecarboxamide(320 mg, 0.84 mmol) was dissolved in a mixture composed of anhydrous DMF(0.5 mL) and anhydrous THF (5 mL) under N₂. NaH (60% in mineral oil, 120mg, 3.0 mmol) was added at room temperature. After 30 min of stirring,the reaction mixture was cooled to −15° C. before a solution ofepichlorohydrin (79 μL, 1.0 mmol) in anhydrous DMF (1 mL) was addeddropwise. The reaction mixture was stirred for 15 min at −15° C., thenfor 8 h at room temperature. MeOH (1 mL) was added and the mixture washeated for 10 min at 105° C. in the microwave oven. The mixture wascooled, filtered and purified by preparative HPLC to give1-(benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-1-(2-hydroxy-3-methoxy-propyl)-1H-indol-5-yl)cyclopropanecarboxamide.¹H NMR (400 MHz, DMSO-d6) δ 8.44 (s, 1H), 7.59 (d, J=1.9 Hz, 1H), 7.31(d, J=8.9 Hz, 1H), 7.03 (dd, J=8.7, 1.9 Hz, 2H), 6.95 (dd, J=8.0, 1.7Hz, 1H), 6.90 (d, J=8.0 Hz, 1H), 6.16 (s, 1H), 6.03 (s, 2H), 4.33 (dd,J=15.0, 4.0 Hz, 1H), 4.19 (dd, J=15.0, 8.1 Hz, 1H), 4.02 (ddd, J=8.7,4.8 Hz, 1H), 3.41-3.32 (m, 2H), 3.30 (s, 3H), 1.41 (s, 9H), 1.41-1.38(m, 2H), 1.03 (dd, J=6.7, 4.0 Hz, 2H). MS (ESI) m/e (M+H⁺) 465.0.

Example 97:1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-1-(2-hydroxy-3-(methyl-amino)propyl)-1H-indol-5-yl)cyclopropanecarboxamide

1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-1H-indol-5-yl)cyclopropanecarboxamide(320 mg, 0.84 mmol) was dissolved in a mixture composed of anhydrous DMF(0.5 mL) and anhydrous THF (5 mL) under N₂. NaH (60% in mineral oil, 120mg, 3.0 mmol) was added at room temperature. After 30 min of stirring,the reaction mixture was cooled to −15° C. before a solution ofepichlorohydrin (79 μL, 1.0 mmol) in anhydrous DMF (1 mL) was addeddropwise. The reaction mixture was stirred for 15 min at −15° C., thenfor 8 h at room temperature. MeNH₂ (2.0 M in MeOH, 1.0 mL) was added andthe mixture was heated for 10 min at 105° C. in the microwave oven. Themixture was cooled, filtered and purified by preparative HPLC to give1-(benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-1-(2-hydroxy-3-(methylamino)propyl)-1H-indol-5-yl)cyclopropanecarboxamide.¹H NMR (400 MHz, DMSO-d6) δ 8.50 (s, 1H), 7.60-7.59 (m, 1H), 7.35 (dd,J=14.3, 8.9 Hz, 1H), 7.10 (d, J=8.8 Hz, 1H), 1H), 6.94 (dd, J=8.0, 1.6Hz, 1H), 6.91 (d, J=7.9 Hz, 1H), 6.20 (d, J=2.3 Hz, 1H), 6.03 (s, 2H),2.82 (d, J=4.7 Hz, 1H), 2.72 (d, J=4.7 Hz, 1H), 2.55 (dd, J=5.2, 5.2 Hz,1H), 2.50 (s, 3H), 1.43 (s, 9H), 1.39 (dd, J=6.4, 3.7 Hz, 2H), 1.04 (dd,J=6.5, 3.9 Hz, 2H). MS (ESI) m/e (M+H⁺) 464.0.

Example 98:(S)-N-(1-(3-Amino-2-hydroxypropyl)-2-tert-butyl-1H-indol-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide

(R)-3-(2-tert-Butyl-5-(1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarbox-amido)-1H-indol-1-yl)-2-hydroxypropyl-4-methylbenzenesulfonate

To a stirred solution of(R)-N-(2-tert-butyl-1-(2,3-dihydroxypropyl)-1H-indol-5-yl)-1-(2,2-difluoro-benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide(3.0 g, 6.1 mmol) in dichloromethane (20 mL) was added triethylamine (2mL) and para-toluenesulfonylchloride (1.3 g, 7.0 mmol). After 18 hours,the reaction mixture was partitioned between 10 mL of water and 10 mL ofethyl acetate. The organic layer was dried over magnesium sulfate,filtered and evaporated. The residue was purified using columnchromatography on silica gel (0-60% ethyl acetate/hexane) providing(R)-3-(2-tert-butyl-5-(1-(2,2-difluorobenzo[d][1,3]-dioxol-5-yl)cyclopropanecarboxamido)-1H-indol-1-yl)-2-hydroxypropyl-4-methyl-benzenesulfonate(3.21 g, 86%). LC/MS (M+1)=641.2. ¹H NMR (400 MHz, CDCl₃) δ 7.77 (d, 2H,J=16 Hz), 7.55 (d, 1H, J=2 Hz), 7.35 (d, 2H, J=16 Hz), 7.31 (m, 3H),6.96 (s, 1H), 6.94 (dd, 1H, J=2, 8 Hz), 6.22 (s, 1H), 4.33 (m, 1H), 4.31(dd, 1H, J=6, 15 Hz), 4.28 (dd, 1H, J=11, 15 Hz), 4.18 (m, 1H), 3.40(dd, 1H, J=3, 6 Hz), 3.36 (dd, 1H, J=3, 6 Hz), 2.46 (s, 3H), 2.40 (br s,1H), 1.74 (m, 2H), 1.40 (s, 9H), 1.11 (m, 2H).

(R)-N-(1-(3-Azido-2-hydroxypropyl)-2-tert-butyl-1H-indol-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide

To a stirred solution(R)-3-(2-tert-butyl-5-(1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1H-indol-1-yl)-2-hydroxypropyl-4-methylbenzenesulfonate(3.2 g, 5.0 mmol) in DMF (6 mL) was added sodium azide (2.0 g, 30 mmol).The reaction was heated at 80° C. for 2 h. The mixture was partitionedbetween 20 mL ethyl acetate and 20 mL water. The layers were separatedand the organic layer was evaporated. The residue was purified usingcolumn chromatography (0-85% ethyl acetate/hexane) to give(R)-N-(1-(3-azido-2-hydroxypropyl)-2-tert-butyl-1H-indol-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)-cyclopropanecarboxamide(2.48 g). LC/MS (M+1)=512.5. ¹H NMR (400 MHz, CDCl₃) δ 7.55 (d, 1H, J=2Hz), 7.31 (m, 3H), 6.96 (s, 1H), 6.94 (dd, 1H, J=2, 8 Hz), 6.22 (s, 1H),4.33 (m, 1H), 4.31 (dd, 1H, J=6, 15 Hz), 4.28 (dd, 1H, J=11, 15 Hz),4.18 (m, 1H), 3.40 (dd, 1H, J=3, 6 Hz), 3.36 (dd, 1H, J=3, 6 Hz), 2.40(br s, 1H), 1.74 (m, 2H), 1.40 (s, 9H), 1.11 (m, 2H).

(S)-N-(1-(3-Amino-2-hydroxypropyl)-2-tert-butyl-1H-indol-5-yl)-1-(2,2-difluoro-benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide

To a stirred solution(R)-N-(1-(3-azido-2-hydroxypropyl)-2-tert-butyl-1H-indol-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide(2.4 g, 4.0 mmol) in MeOH (25 mL) was added 5% Pd/C (2.4 g) under aHydrogen gas filled balloon. After 18 h, the reaction mixture wasfiltered through celite and rinsed with 300 mL ethyl acetate. Theorganic layer was washed with 1 N HCl and evaporated to give(S)-N-(1-(3-amino-2-hydroxypropyl)-2-tert-butyl-1H-indol-5-yl)-1-(2,2-difluoro-benzo[d][1,3]-dioxol-5-yl)cyclopropane-carboxamide(1.37 g). MS (M+1)=486.5.

Example 99: (S)-Methyl3-(2-tert-butyl-5-(1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1H-indol-1-yl)-2-hydroxypropylcarbamate

To a stirred solution(R)-N-(1-(3-amino-2-hydroxypropyl)-2-tert-butyl-1H-indol-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide(0.10 g, 0.20 mmol) in methanol (1 mL) was added 2 drops oftriethylamine and methylchloroformyl chloride (0.020 mL, 0.25 mmol).After 30 min, the reaction mixture was filtered and purified usingreverse phase HPLC providing (S)-methyl3-(2-tert-butyl-5-(1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclo-propanecarboxamido)-1H-indol-1-yl)-2-hydroxypropylcarbamate.The retention time on a three minute run is 1.40 min. LC/MS (M+1)=544.3.¹H NMR (400 MHz, CDCl₃) δ 7.52 (d, 1H, J=2 Hz), 7.30 (dd, 1H, J=2, 8Hz), 7.28 (m, 1H), 7.22 (d, 1H, J=8 Hz), 7.14 (d, 1H, J=8 Hz), 7.04 (brs, 1H), 6.97 (dd, 1H, J=2, 8 Hz), 6.24 (s, 1H), 5.19 (1H, br s), 4.31(dd, 1H, J=6, 15 Hz), 4.28 (dd, 1H, J=11, 15 Hz), 4.18 (m, 1H), 3.70 (s,3H), 3.40 (dd, 1H, J=3, 6 Hz), 3.36 (dd, 1H, J=3, 6 Hz), 3.26 (m, 1H),1.74 (m, 2H), 1.40 (s, 9H), 1.11 (m, 2H).

Example 100:4-(5-(1-(Benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-2-tert-butyl-1H-indol-1-yl)butanoicAcid

1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butylindolin-5-yl)cyclopropanecarboxamide

To a solution of1-(benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-1H-indol-5-yl)cyclo-propanecarboxamide(851 mg, 2.26 mmol) in acetic acid (60 mL) was added NaBH₃CN (309 mg,4.91 mmol) at 0° C. The reaction mixture was stirred for 5 min at roomtemperature after which no starting material could be detected by LCMS.The solvent was evaporated under reduced pressure and the residue waspurified by column chromatography on silica gel (5-40% ethylacetate/hexanes) to give1-(benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butylindolin-5-yl)cyclopropanecarboxamide(760 mg, 89%).

4-(5-(1-(Benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-2-tert-butylindolin-1-yl)butanoicAcid

To a solution of1-(benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butylindolin-5-yl)cyclopropanecarboxamide(350 mg, 0.93 mmol, 1 eq) in anhydrous methanol (6.5 mL) and AcOH (65μL) was added 4-oxobutanoic acid (15% in water, 710 mg, 1.0 mmol) atroom temperature. After 20 min of stirring, NaBH₃CN (130 mg, 2.0 mmol)was added in one portion and the reaction mixture was stirred foranother 4 h at room temperature. The reaction mixture was quenched byaddition of AcOH (0.5 mL) at 0° C. and the solvent was removed underreduced pressure. The residue was purified by column chromatography onsilica gel (5-75% ethyl acetate/hexanes) to give4-(5-(1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-2-tert-butylindolin-1-yl)butanoicacid (130 mg, 30%).

4-(5-(1-(Benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-2-tert-butyl-1H-indol-1-yl)butanoicAcid

4-(5-(1-(Benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-2-tert-butylindolin-1-yl)butanoicacid (130 mg, 0.28 mmol) was taken up in a mixture ofacetonitrile-H₂O-TFA. The solvent was removed under reduced pressure andthe residue obtained was dissolved in CDCl₃. After a brief exposition todaylight (5-10 min), the solution turned purple. The mixture was stirredopen to the atmosphere at room temperature until complete disappearanceof the starting material (8 h). Solvent was removed under reducedpressure and the residue was purified by reverse phrase HPLC to give4-(5-(1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-2-tert-butyl-1H-indol-1-yl)butanoicacid. ¹H NMR (400 MHz, CDCl₃) δ 7.52 (d, J=1.9 Hz, 1H), 7.18 (d, J=2.1Hz, 1H), 7.16 (s, 1H), 7.03 (dd, J=9.4, 1.9 Hz, 1H), 7.00-6.98 (m, 2H),6.85 (d, J=7.9 Hz, 1H), 6.16 (s, 1H), 6.02 (s, 2H), 4.29-4.24 (m, 2H),2.48 (dd, J=6.9, 6.9 Hz, 2H), 2.12-2.04 (m, 2H), 1.69 (dd, J=6.8, 3.7Hz, 2H), 1.43 (s, 9H), 1.09 (dd, J=6.8, 3.7 Hz, 2H). MS (ESI) m/e (M+H⁺)463.0.

Example 101:1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-1-(4-(2-hydroxyethyl-amino)-4-oxobutyl)-1H-indol-5-yl)cyclopropanecarboxamide

To a solution of4-(5-(1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-2-tert-butyl-1H-indol-1-yl)butanoicacid (10 mg) in anhydrous DMF (0.25 mL) were successively added Et₃N(9.5 mL, 0.069 mmol) and HBTU (8.2 mg, 0.022 mmol). After stirring for10 min at 60° C., ethanolamine (1.3 μL, 0.022 mmol) was added, and themixture was stirred for another 4 h at 60° C.1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-1-(4-(2-hydroxyethyl-amino)-4-oxobutyl)-1H-indol-5-yl)cyclopropanecarboxamide(5.8 mg, 64%) was obtained after purification by preparative HPLC. MS(ESI) m/e (M+H⁺) 506.0.

Example 102:1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-1-(2-(dimethylamino)-2-oxoethyl)-1H-indol-5-yl)cyclopropanecarboxamide

To a solution of1-(benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butylindolin-5-yl)cyclopropanecarboxamide(62 mg, 0.16 mmol) in anhydrous DMF (0.11 mL) and THF (1 mL) was addedNaH (60% in mineral oil, 21 mg, 0.51 mmol) at room temperature under N₂.After 30 min of stirring, the reaction mixture was cooled to 0° C. and2-chloro-N,N-dimethylacetamide (11 mL, 0.14 mmol,) was added. Thereaction mixture was stirred for 5 min at 0° C. and then for 10 h atroom temperature. The mixture was purified by preparative HPLC and theresultant solid was dissolved in DMF (0.6 mL) in the presence of Pd-C(10 mg). The mixture was stirred open to the atmosphere overnight atroom temperature. The reaction mixture was filtrated and purified bypreparative HPLC providing 1-(benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-1-(2-(dimethylamino)-2-oxoethyl)-1H-indol-5-yl)cyclopropanecarboxamide.MS (ESI) m/e (M+H⁺) 462.0.

Example 103:3-(2-tert-Butyl-5-(1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclo-propanecarboxamido)-1H-indol-1-yl)propanoicAcid

N-(2-tert-Butyl-1-(2-chloroethyl)indolin-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide

To a solution ofN-(2-tert-butyl-1-(2-cyanoethyl)indolin-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide(71 mg, 0.17 mmol) in anhydrous dichloromethane (1 mL) was addedchloroacetaldehyde (53 μL, 0.41 mmol) at room temperature under N₂.After 20 min of stirring, NaBH(OAc)₃ (90 mg, 0.42 mmol) was added in twoportions. The reaction mixture was stirred overnight at roomtemperature. The product was purified by column chromatography on silicagel (2-15% ethyl acetate/hexanes) providingN-(2-tert-butyl-1-(2-chloroethyl)indolin-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide(51 mg, 63%).

N-(2-tert-Butyl-1-(2-cyanoethyl)indolin-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide

N-(2-tert-butyl-1-(2-chloroethyl)indolin-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide(51 mg), NaCN (16 mg, 0.32 mmol) and KI (cat) in EtOH (0.6 mL) and water(0.3 mL) were combined and heated at 110° C. for 30 min in themicrowave. The solvent was removed under reduced pressure and theresidue was purified by column chromatography on silica gel (2-15% ethylacetate/hexanes) providingN-(2-tert-butyl-1-(2-cyanoethyl)indolin-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide(24 mg, 48%).

3-(2-tert-Butyl-5-(1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclo-propanecarbox-amido)-1H-indol-1-yl)propanoicacid

N-(2-tert-butyl-1-(2-cyanoethyl)indolin-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropane-carboxamide(24 mg, 0.050 mmol) was taken up in 50% aq. KOH (0.5 mL) and 1,4-dioxane(1 mL). The mixture was heated at 125° C. for 2 h. The solvent wasremoved and the residue was purified by preparative HPLC. The residuewas dissolved in CDCl₃ (1 mL) then briefly exposed to daylight. Thepurple solution that formed was stirred until complete disappearance ofthe starting material (1 h). The solvent was removed under reducedpressure and the residue was purified by preparative HPLC providing3-(2-tert-butyl-5-(1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclo-propanecarboxamido)-1H-indol-1-yl)propanoicacid. MS (ESI) m/e (M+H⁺) 485.0.

Example 104:1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-6-fluoro-1-(2-hydroxy-ethyl)-1H-indol-5-yl)cyclopropenecarboxamide

To a solution of1-(benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-6-fluoroindolin-5-yl)cyclopropanecarboxamide(340 mg, 0.86 mmol) in anhydrous MeOH (5.7 mL) containing 1% of aceticacid was added glyoxal 40% in water (0.60 mL, 5.2 mmol) at roomtemperature under N₂. After 20 min of stirring, NaBH₃CN (120 mg, 1.9mmol) was added in one portion and the reaction mixture was stirredovernight at room temperature. The solvent was removed under reducedpressure and the residue obtained was purified by column chromatographyon silica gel (10-40% ethyl acetate/hexanes) providing a pale yellow oilwhich was treated with 50/50 CH₃CN—H₂O containing 0.05% TFA and CDCl₃.Solvent was removed under reduced pressure and the residue was purifiedby column chromatography on silica gel (20-35% ethyl acetate/hexanes) togive1-(benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-6-fluoro-1-(2-hydroxyethyl)-1H-indol-5-yl)cyclopropanecarboxamide.¹H NMR (400 MHz, CDCl₃) δ 8.02 (d, J=7.7 Hz, 1H), 7.30 (d, J=2.1 Hz,1H), 6.93 (dd, J=1.6, 7.9 Hz, 1H), 6.90 (d, J=1.6 Hz, 1H), 6.90 (d,J=1.6 Hz, 1H), 6.78 (d, J=7.9 Hz, 1H), 6.08 (s, 1H), 5.92 (s, 2H), 4.21(dd, J=6.9, 6.9 Hz, 2H), 3.68 (m, 2H), 2.28 (s, 1H), 1.60 (dd, J=3.7,6.7 Hz, 2H), 1.35-1.32 (m, 9H), 1.04 (dd, J=3.7, 6.8 Hz, 2H). MS (ESI)m/e (M+H⁺) 439.0.

Example 105:1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-6-fluoro-1-(3-hydroxy-propyl)-1H-indol-5-yl)cyclopropanecarboxamide

3-(Benzyloxy)propanal

To a suspension of PCC (606 mg, 2.82 mmol) in anhydrous dichloromethane(8 mL) at room temperature under N₂ was added a solution of3-benzyloxy-1-propanol (310 mg, 1.88 mmol) in anhydrous dichloromethane.The reaction mixture was stirred overnight at room temperature,filtrated through Celite, and concentrated. The residue was purified bycolumn chromatography on silica gel (1-10% ethyl acetate/hexanes) togive 3-(benzyloxy)propanal (243 mg, 79%).

1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-6-fluoro-1-(3-hydroxypropyl)-1H-indol-5-yl)cyclopropanecarboxamide

To a solution of1-(benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-6-fluoroindolin-5-yl)cyclopropanecarboxamide(160 mg, 0.50 mmol) in anhydrous dichloromethane (3.4 mL) was added3-(benzyloxy)propanal (160 mg, 0.98 mmol) at room temperature. After 10min of stirring, NaBH(OAc)₃ (140 mg, 0.65 mmol) was added in one portionand the reaction mixture was stirred for 4 h at room temperature. Thesolvent was removed under reduced pressure and the residue was taken-upin a mixture of 50/50 CH₃CN—H₂O containing 0.05% TFA. The mixture wasconcentrated to dryness and the residue was dissolved in CDCl₃ (5 mL)and briefly exposed to daylight. The purple solution was stirred open tothe atmosphere at room temperature for 2 h. The solvent was removedunder reduced pressure and the residue was treated with Pd-C (10 mg) inMeOH (2 mL) under 1 atm of H₂ for 2 h. The catalyst was filtered throughCelite and the solvent was removed under reduced pressure. The residuewas purified by preparative TLC 30% ethyl acetate/hexanes to provide1-(benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-6-fluoro-1-(3-hydroxypropyl)-1H-indol-5-yl)cyclopropanecarboxamide(18 mg, 8% from1-(benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-6-fluoroindolin-5-yl)cyclopropane-carboxamide).¹H NMR (400 MHz, CDCl₃) δ 8.11 (d, J=7.8 Hz, 1H), 7.31 (d, J=2.2 Hz,1H), 6.94 (dd, J=7.9, 1.7 Hz, 1H), 6.91 (d, J=1.6 Hz, 1H), 6.85 (d,J=11.7 Hz, 1H), 6.79 (d, J=7.9 Hz, 1H), 6.10 (s, 1H), 5.94 (s, 2H),4.25-4.21 (m, 2H), 3.70 (dd, J=5.7, 5.7 Hz, 2H), 1.93-1.86 (m, 2H), 1.61(dd, J=6.8, 3.7 Hz, 2H), 1.35 (s, 9H), 1.04 (dd, J=6.8, 3.7 Hz, 2H). MS(ESI) m/e (M+H⁺) 453.0.

Example 106:N-(1-(2-Acetamidoethyl)-2-tert-butyl-1H-indol-5-yl)-1-(benzo[d][1,3]-dioxol-5-yl)cyclopropanecarboxamide

N-(1-(2-azidoethyl)-2-tert-butyl-1H-indol-5-yl)-1-(benzo[d][1,3]dioxol-5-yl)-cyclopropanecarboxamide

To a solution of1-(benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butylindolin-5-yl)cyclopropane-carboxamide(73 mg, 0.19 mmol) in anhydrous dichloromethane (1.2 mL) was addedchloroacetaldehyde (60 μL, 0.24 mmol) at room temperature. After 10 minof stirring, NaBH(OAc)₃ (52 mg, 0.24 mmol) was added in one portion andthe reaction mixture was stirred for another 30 min at room temperature.The solvent was removed under reduced pressure and the residue waspurified by preparative HPLC to give the indoline, which oxidized to thecorresponding indole when taken-up in CDCl₃. The resulting indole wastreated with NaN₃ (58 mg, 0.89 mmol) and NaI (cat) in anhydrous DMF (0.8mL) for 2 h at 85° C. The reaction mixture was purified by preparativeHPLC providingN-(1-(2-azidoethyl)-2-tert-butyl-1H-indol-5-yl)-1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide(15 mg, 18% from1-(benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butylindolin-5-yl)cyclopropane-carboxamide).

N-(1-(2-Acetamidoethyl)-2-tert-butyl-1H-indol-5-yl)-1-(benzo[d][1,3]-dioxol-5-yl)cyclopropanecarboxamide

A solution ofN-(1-(2-azidoethyl)-2-tert-butyl-1H-indol-5-yl)-1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide(13 mg, 0.029 mmol) in MeOH—AcOH (0.2 mL, 99:1) in the presence of Pd-C(2 mg) was stirred at room temperature under 1 atm of H₂ for 2 h,filtered through Celite, and concentrated under reduced pressure. Thecrude product was treated with AcCl (0.05 mL) and Et₃N (0.05 mL) inanhydrous THF (0.2 mL) at 0° C. for 30 min and then 1 h at roomtemperature. The mixture was purified by preparative HPLC providingN-(1-(2-acetamidoethyl)-2-tert-butyl-1H-indol-5-yl)-1-(benzo[d][1,3]-dioxol-5-yl)cyclopropanecarboxamide.MS (ESI) m/e (M+H⁺) 462.0.

Example 107:N-(2-tert-Butyl-1-(3-cyano-2-hydroxypropyl)-1H-indol-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide

3-(2-tert-Butyl-5-(1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarbox-amido)-1H-indol-1-yl)-2-hydroxypropyl-4-methylbenzenesulfonate

To a solution ofN-(2-tert-butyl-1-(2,3-dihydroxypropyl)-1H-indol-5-yl)-1-(2,2-difluorobenzo[d][1,3]-dioxol-5-yl)cyclopropanecarboxamide(172 mg, 0.35 mmol) in anhydrous dichloromethane (1.4 mL) at 0° C. inthe presence of Et₃N (56 μL, 0.40 mmol) was added TsCl (71 mg, 0.37mmol). The reaction mixture was stirred for 2 h at room temperaturebefore being cooled to 0° C. and another portion of TsCl (71 mg, 0.37mmol) was added. After 1 h of stirring at room temperature, the mixturewas purified by column chromatography on silica gel (10-30% ethylacetate/hexanes) providing3-(2-tert-butyl-5-(1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1H-indol-1-yl)-2-hydroxypropyl-4-methylbenzene-sulfonate(146 mg, 64%).

N-(2-tert-Butyl-1-(3-cyano-2-hydroxypropyl)-1H-indol-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide

N-(2-tert-Butyl-1-(3-cyano-2-hydroxypropyl)-1H-indol-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)-cyclopropanecarboxamide(145 mg, 0.226 mmol) was treated with powdered NaCN (34 mg, 0.69 mmol)in anhydrous DMF (1.5 mL) at 85° C. for 2 h. The reaction mixture wascooled down to room temperature before it was diluted withdichloromethane (10 mL) and aq. sat. NaHCO₃ (10 mL). The organic phasewas separated and the aqueous phase was extracted with dichloromethane(2×10 mL). The organic phases were combined, washed with brine, driedwith sodium sulfate, filtered then concentrated. The residue waspurified by column chromatography on silica gel (25-55% ethylacetate/hexanes) providingN-(2-tert-butyl-1-(3-cyano-2-hydroxypropyl)-1H-indol-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide(89 mg, 79%). ¹H NMR (400 MHz, CDCl₃) δ 7.43 (d, J=1.9 Hz, 1H),7.20-7.16 (m, 2H), 7.08 (d, J=8.8 Hz, 1H), 7.04 (d, J=8.2 Hz, 1H), 6.94(s, 1H), 6.88 (dd, J=8.7, 2.0 Hz, 1H), 6.16 (s, 1H), 4.32-4.19 (m, 3H),2.83 (s, 1H), 2.40 (dd, J=5.2, 5.2 Hz, 2H), 1.62 (dd, J=6.6, 3.6 Hz,2H), 1.35 (s, 9H), 1.04 (dd, J=6.9, 3.9 Hz, 2H). MS (ESI) m/e (M+H⁺)496.0.

Example 108:N-(2-tert-Butyl-1-(2-hydroxy-3-(2H-tetrazol-5-yl)propyl)-1H-indol-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide

To a solution ofN-(2-tert-butyl-1-(3-cyano-2-hydroxypropyl)-1H-indol-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide(27 mg, 0.054 mmol) in anhydrous DMF (1.2 mL) were successively addedNH₄Cl (35 mg, 0.65 mmol) and NaN₃ (43 mg, 0.65 mmol) at roomtemperature. The reaction mixture was stirred for 4 h at 110° C. in themicrowave, at which stage 50% of the starting material was converted tothe desired product. The reaction mixture was purified by preparativeHPLC to provideN-(2-tert-butyl-1-(2-hydroxy-3-(2H-tetrazol-5-yl)propyl)-1H-indol-5-yl)-1-(2,2-difluorobenzo-[d][1,3]dioxol-5-yl)cyclopropanecarboxamide.MS (ESI) m/e (M+H⁺) 539.0.

Example 109:4-(2-tert-Butyl-5-(1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclo-propanecarboxamido)-1H-indol-1-yl)-3-hydroxybutanoicAcid

A solution ofN-(2-tert-butyl-1-(3-cyano-2-hydroxypropyl)-1H-indol-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide(14 mg, 0.028 mmol) in methanol (0.8 mL) and 4 M NaOH (0.8 mL) wasstirred at 60° C. for 4 h. The reaction mixture was neutralized with 4 MHCl and concentrated. The residue was purified by preparative HPLC toprovide4-(2-tert-butyl-5-(1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1H-indol-1-yl)-3-hydroxybutanoicacid. MS (ESI) m/e (M+H⁺) 515.0.

Example 110:N-(1-(2-(2H-Tetrazol-5-yl)ethyl)-2-tert-butyl-1H-indol-5-yl)-1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide

1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-1-(2-cyanoethyl)indolin-5-yl)-cyclopropanecarboxamide

To a solution of1-(benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-1-(2-chloroethyl)indolin-5-yl)cyclopropanecarboxamide(66 mg, 0.15 mmol) in ethanol (0.8 mL) and water (0.4 mL) were addedNaCN (22 mg, 0.45 mmol) and KI (cat) at room temperature. The reactionmixture was stirred for 30 min at 110° C. in the microwave before beingpurified by column chromatography on silica gel (5-15% ethylacetate/hexanes) to provide1-(benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-1-(2-cyano-ethyl)indolin-5-yl)cyclopropanecarboxamide(50 mg, 77%).

N-(1-(2-(2H-Tetrazol-5-yl)ethyl)-2-tert-butyl-1H-indol-5-yl)-1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide

To a solution of1-(benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-1-(2-cyano-ethyl)indolin-5-yl)cyclopropanecarboxamide(50 mg, 0.12 mmol) in anhydrous DMF (2.6 mL) was added NH₄Cl (230 mg,4.3 mmol) and NaN₃ (280 mg, 4.3 mmol). The reaction mixture was stirredfor 30 min at 110° C. in the microwave, filtrated, and purified bypreparative HPLC. The solid residue was dissolved in CDCl₃ (3 mL) andbriefly (2 to 4 min) exposed to daylight, which initiated a color change(purple). After 2 h of stirring open to the atmosphere at roomtemperature, the solvent was removed and the residue was purified bypreparative HPLC to giveN-(1-(2-(2H-tetrazol-5-yl)ethyl)-2-tert-butyl-1H-indol-5-yl)-1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide.MS (ESI) m/e (M+H⁺) 473.0.

Example 111:1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-6-fluoro-1-((tetrahydro-2H-pyran-3-yl)methyl)-1H-indol-5-yl)cyclopropanecarboxamide

To a solution of1-(benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-6-fluoroindolin-5-yl)cyclopropane-carboxamide(150 mg, 0.38 mmol) in anhydrous dichloromethane (2.3 mL) at roomtemperature under N₂ was added tetrahydropyran-3-carbaldehyde (54 mg,0.47 mmol). After 20 min of stirring, NaBH(OAc)₃ (110 mg, 0.51 mmol) wasadded in one portion at room temperature. The reaction mixture wasstirred for 6 h at room temperature before being purified by columnchromatography on silica gel (5-20% ethyl acetate/hexanes) to provide1-(benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-6-fluoro-1-((tetrahydro-2H-pyran-3-yl)methyl)indolin-5-yl)cyclopropanecarboxamide(95 mg, 50%). CDCl₃ was added to the indoline and the solution wasallowed to stir overnight at ambient temperature. The solution wasconcentrated to give1-(benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-6-fluoro-1-((tetrahydro-2H-pyran-3-yl)methyl)-1H-indol-5-yl)cyclopropanecarboxamide.MS (ESI) m/e (M+H⁺) 493.0.

Example 112:1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-(2-hydroxypropan-2-yl)-1H-indol-5-yl)cyclopropanecarboxamide

Methyl5-(1-(benzo[d][1,3]dioxol-5-yl)cyclopropane-carboxamido)-1H-indole-2-carboxylate(100 mg, 0.255 mmol) was dissolved in anhydrous tetrahydrofuran (2 mL)under an argon atmosphere. The solution was cooled to 0° C. in an icewater bath before methyllithium (0.85 mL, 1.6 M in diethyl ether) wasadded by syringe. The mixture was allowed to warm to room temperature.The crude product was then partitioned between a saturated aqueoussolution of sodium chloride (5 mL) and dichloromethane (5 mL). Theorganic layers were combined, dried over sodium sulfate, filtered,evaporated to dryness, and purified on 12 g of silica gel utilizing agradient of 20-80% ethyl acetate in hexanes to yield1-(benzo[d][1,3]dioxol-5-yl)-N-(2-(2-hydroxypropan-2-yl)-1H-indol-5-yl)cyclopropanecarboxamide(35 mg, 36%) as a white solid. ESI-MS m/z calc. 378.2, found 379.1(M+1)⁺. Retention time of 2.18 minutes. ¹H NMR (400 MHz, DMSO-d6) δ10.78 (s, 1H), 8.39 (s, 1H), 7.57 (d, J=1.7 Hz, 1H), 7.17 (d, J=8.6 Hz,1H), 7.03-6.90 (m, 4H), 6.12 (d, J=1.5 Hz, 1H), 6.03 (s, 2H), 5.18 (s,1H), 1.50 (s, 6H), 1.41-1.38 (m, 2H), 1.05-0.97 (m, 2H).

Example 113:N-(2-(1-Amino-2-methylpropan-2-yl)-1H-indol-5-yl)-1-(benzo[d][1,3]-dioxol-5-yl)cyclopropanecarboxamide

Trifluoroacetic acid (0.75 mL) was added to a solution of tert-butyl2-(5-(1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1H-indol-2-yl)-2-methylpropylcarbamate(77 mg, 0.16 mmol) in dichloromethane (3 mL) and the mixture was stirredat room temperature for 1.5 h. The mixture was evaporated, dissolved indichloromethane, washed with saturated sodium bicarbonate solution,dried over magnesium sulfate and evaporated to dryness to giveN-(2-(1-amino-2-methylpropan-2-yl)-1H-indol-5-yl)-1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide(53 mg, 86%). ¹H NMR (400 MHz, CDCl₃) δ 9.58 (s, 1H), 7.60 (d, J=1.6 Hz,1H), 7.18-7.15 (m, 2H), 7.02-6.94 (m, 3H), 6.85 (d, J=7.8 Hz, 1H), 6.14(d, J=1.2 Hz, 1H), 6.02 (s, 2H), 2.84 (s, 2H), 1.68 (dd, J=3.6, 6.7 Hz,2H), 1.32 (s, 6H), 1.08 (dd, J=3.7, 6.8 Hz, 2H).

Example 114:1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-(1-(dimethylamino)-2-methyl-propan-2-yl)-1H-indol-5-yl)cyclopropanecarboxamide

To a solution ofN-(2-(1-amino-2-methylpropan-2-yl)-1H-indol-5-yl)-1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide(20 mg, 0.051 mmol) in DMF (1 mL) was added potassium carbonate (35 mg,0.26 mmol) and iodomethane (7.0 μL, 0.11 mmol). The mixture was stirredfor 2 h. Water was added and the mixture was extracted withdichloromethane. Combined organic phases were dried over magnesiumsulfate, evaporated, coevaporated with toluene (3×) and purified bysilica gel chromatography (0-30% EtOAc in hexane) to give1-(benzo[d][1,3]dioxol-5-yl)-N-(2-(1-(dimethylamino)-2-methylpropan-2-yl)-1H-indol-5-yl)cyclopropanecarboxamide(7 mg, 33%). ¹H NMR (400 MHz, CDCl₃) δ 9.74 (s, 1H), 7.58 (d, J=1.9 Hz,1H), 7.20 (d, J=8.6 Hz, 1H), 7.15 (s, 1H), 7.01-6.95 (m, 3H), 6.85 (d,J=7.9 Hz, 1H), 6.10 (d, J=0.9 Hz, 1H), 6.02 (s, 2H), 2.43 (s, 2H), 2.24(s, 6H), 1.68 (dd, J=3.7, 6.7 Hz, 2H), 1.33 (s, 6H), 1.08 (dd, J=3.7,6.8 Hz, 2H).

Example 115:N-(2-(1-Acetamido-2-methylpropan-2-yl)-1H-indol-5-yl)-1-(benzo[d][1,3]-dioxol-5-yl)cyclopropanecarboxamide

To a solution ofN-(2-(1-amino-2-methylpropan-2-yl)-1H-indol-5-yl)-1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide(21 mg, 0.054 mmol) in dichloromethane (1 mL) was added pyridine (14 μL,0.16 mmol) followed by acetic anhydride (6.0 μL, 0.059 mmol). Themixture was stirred for 2 h. Water was added and the mixture wasextracted with dichloromethane, evaporated, coevaporated with toluene(3×) and purified by silica gel chromatography (60-100% ethylacetate inhexane) to giveN-(2-(1-acetamido-2-methylpropan-2-yl)-1H-indol-5-yl)-1-(benzo[d][1,3]-dioxol-5-yl)cyclopropanecarboxamide(17 mg, 73%). ¹H NMR (400 MHz, DMSO) δ 10.79 (s, 1H), 8.39 (s, 1H), 7.66(t, J=6.2 Hz, 1H), 7.56 (d, J=1.7 Hz, 1H), 7.18-7.14 (m, 1H), 7.02-6.89(m, 4H), 6.08 (d, J=1.5 Hz, 1H), 6.03 (s, 2H), 3.31 (d, J=6.2 Hz, 2H),1.80 (s, 3H), 1.41-1.38 (m, 2H), 1.26 (s, 6H), 1.04-1.01 (m, 2H).

Example 116:1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-(2-methyl-4-(1H-tetrazol-5-yl)butan-2-yl)-1H-indol-5-yl)cyclopropanecarboxamide

1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-(4-cyano-2-methylbutan-2-yl)-1H-indol-5-yl)cyclopropanecarboxamide(83 mg, 0.20 mmol) was dissolved in N,N-dimethylformamide (1 mL)containing ammonium chloride (128 mg, 2.41 mmol), sodium azide (156 mg,2.40 mmol), and a magnetic stir bar. The reaction mixture was heated at110° C. for 40 minutes in a microwave reactor. The crude product wasfiltered and then purified by preparative HPLC using a gradient of 0-99%acetonitrile in water containing 0.05% trifluoroacetic acid to yield1-(benzo[d][1,3]dioxol-5-yl)-N-(2-(2-methyl-4-(1H-tetrazol-5-yl)butan-2-yl)-1H-indol-5-yl)cyclopropanecarboxamide.ESI-MS m/z calc. 458.2, found 459.2 (M+1)⁺. Retention time of 1.53minutes. ¹H NMR (400 MHz, CD₃CN) 9.23 (s, 1H), 7.51-7.48 (m, 2H), 7.19(d, J=8.6 Hz, 1H), 7.06-7.03 (m, 2H), 6.95-6.89 (m, 2H), 6.17 (dd,J=0.7, 2.2 Hz, 1H), 6.02 (s, 2H), 2.61-2.57 (m, 2H), 2.07-2.03 (m, 2H),1.55-1.51 (m, 2H), 1.39 (s, 6H), 1.12-1.09 (m, 2H).

Example 117:1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-(piperidin-2-yl)-1H-indol-5-yl)cyclopropanecarboxamide

tert-Butyl2-(5-(1-(benzo[d][1,3]dioxol-5-yl)cyclo-propanecarboxamido)-1H-indol-2-yl)piperidine-1-carboxylate(55 mg, 0.11 mmol) was dissolved in dichloromethane (2.5 mL) containingtrifluoroacetic acid (1 mL). The reaction mixture was stirred for 6 h atroom temperature. The crude product was purified by preparative HPLCusing a gradient of 0-99% acetonitrile in water containing 0.05%trifluoroacetic acid to yield1-(benzo[d][1,3]dioxol-5-yl)-N-(2-(piperidin-2-yl)-1H-indol-5-yl)cyclopropanecarboxamide.ESI-MS m/z calc. 403.2, found 404.4 (M+1)⁺. Retention time of 0.95minutes.

Example 118: 5-tert-Butyl-1H-indol-6-ylamine

2-Bromo-4-tert-butyl-phenylamine

To a solution of 4-tert-Butyl-phenylamine (447 g, 3.00 mol) in DMF (500mL) was added dropwise NBS (531 g, 3.00 mol) in DMF (500 mL) at roomtemperature. Upon completion, the reaction mixture was diluted withwater and extracted with EtOAc. The organic layer was washed with water,brine, dried over Na₂SO₄ and concentrated. The crude product wasdirectly used in the next step without further purification.

2-Bromo-4-tert-butyl-5-nitro-phenylamine

2-Bromo-4-tert-butyl-phenylamine (160 g, 0.71 mol) was added dropwise toH₂SO₄ (410 mL) at room temperature to yield a clear solution. This clearsolution was then cooled down to −5 to −10° C. A solution of KNO₃ (83 g,0.82 mol) in H₂SO₄ (410 mL) was added dropwise while the temperature wasmaintained between −5 to −10° C. Upon completion, the reaction mixturewas poured into ice/water and extracted with EtOAc. The combined organiclayers were washed with 5% Na₂CO₃ and brine, dried over Na₂SO₄ andconcentrated. The residue was purified by a column chromatography (ethylacetate/petroleum ether 1:10) to give2-bromo-4-tert-butyl-5-nitro-phenylamine as a yellow solid (150 g, 78%).

4-tert-Butyl-5-nitro-2-trimethylsilanylethynyl-phenylamine

To a mixture of 2-bromo-4-tert-butyl-5-nitro-phenylamine (27.3 g, 100mmol) in toluene (200 mL) and water (100 mL) was added Et₃N (27.9 mL,200 mmol), Pd(PPh₃)₂Cl₂ (2.11 g, 3.00 mmol), CuI (950 mg, 0.500 mmol)and trimethylsilyl acetylene (21.2 mL, 150 mmol) under a nitrogenatmosphere. The reaction mixture was heated at 70° C. in a sealedpressure flask for 2.5 h., cooled down to room temperature and filteredthrough a short plug of Celite. The filter cake was washed with EtOAc.The combined filtrate was washed with 5% NH₄OH solution and water, driedover Na₂SO₄ and concentrated. The crude product was purified by columnchromatography (0-10% ethyl acetate/petroleum ether) to provide4-tert-butyl-5-nitro-2-trimethylsilanylethynyl-phenylamine as a brownviscous liquid (25 g, 81%).

5-tert-Butyl-6-nitro-1H-indole

To a solution of4-tert-butyl-5-nitro-2-trimethylsilanylethynyl-phenylamine (25 g, 86mmol) in DMF (100 mL) was added CuI (8.2 g, 43 mmol) under a nitrogenatmosphere. The mixture was heated at 135° C. in a sealed pressure flaskovernight, cooled down to room temperature and filtered through a shortplug of Celite. The filter cake was washed with EtOAc. The combinedfiltrate was washed with water, dried over Na₂SO₄ and concentrated. Thecrude product was purified by column chromatography (10-20% ethylaetate/hexane) to provide 5-tert-butyl-6-nitro-1H-indole as a yellowsolid (13 g, 69%).

5-tert-Butyl-1H-indol-6-ylamine

Raney Nickel (3 g) was added to 5-tert-butyl-6-nitro-1H-indole (15 g, 67mmol) in methanol (100 mL). The mixture was stirred under hydrogen (1atm) at 30° C. for 3 h. The catalyst was filtered off. The filtrate wasdried over Na₂SO₄ and concentrated. The crude dark brown viscous oil waspurified by column chromatography (10-20% ethyl acetate/petroleum ether)to give 5-tert-butyl-1H-indol-6-ylamine as a gray solid (11 g, 87%). ¹HNMR (300 MHz, DMSO-d6) δ 10.3 (br s, 1H), 7.2 (s, 1H), 6.9 (m, 1H), 6.6(s, 1H), 6.1 (m, 1H), 4.4 (br s, 2H), 1.3 (s, 9H).

A person skilled in the chemical arts can use the examples and schemesalong with known synthetic methodologies to synthesize compounds of thepresent invention, including the compounds in Table 3, below.

TABLE 3 Physical data of exemplary compounds. Compound LC/MS LC/RT No.M + 1 Min NMR 1 373.3 2.49 2 469.4 3.99 3 381.3 3.69 4 448.3 1.75 5389.3 3.3 6 463 1.87 7 363.3 3.7 8 405.5 3.87 9 487.3 2.12 H NMR (400MHz, DMSO- d6) 8.65 (s, 1H), 7.55 (d, J = 1.7 Hz, 1H), 7.49 (d, J = 1.4Hz, 1H), 7.38 (d, J = 8.3 Hz, 1H), 7.30- 7.25 (m, 2H), 7.08 (dd, J =8.8, 1.9 Hz, 1H), 6.11 (s, 1H), 4.31 (t, J = 7.4 Hz, 2H), 3.64 (t, J =7.3 Hz, 2H), 3.20 (t, J = 7.6 Hz, 2H), 1.92 (t, J = 7.6 Hz, 2H), 1.45(m, 2H), 1.39 (s, 6H), 1.10 (m, 2H) 10 388 3.34 11 452.3 2.51 12 5272.36 13 498 1.85 14 404.5 1.18 15 369.2 3.81 16 419.2 2.24 17 389.2 2.02H NMR (400 MHz, DMSO) 8.41 (s, 1H), 7.59 (d, J = 1.8 Hz, 1H), 7.15 (d, J= 8.6 Hz, 1H), 7.06-7.02 (m, 2H), 6.96- 6.90 (m, 2H), 6.03 (s, 2H), 5.98(d, J = 0.7 Hz, 1H), 4.06 (t, J = 6.8 Hz, 2H), 2.35 (t, J = 6.8 Hz, 2H),1.42-1.38 (m, 2H), 1.34 (s, 6H), 1.05-1.01 (m, 2H) 18 395.3 3.6 H NMR(400 MHz, DMSO) 10.91 (s, 1H), 7.99 (s, 1H), 7.67 (d, J = 7.7 Hz, 1H),7.08- 6.92 (m, 4H), 6.09-6.03 (m, 3H), 1.47-1.42 (m, 2H), 1.31 (d, J =7.3 Hz, 9H), 1.09-1.05 (m, 2H) 19 457.2 1.97 H NMR (400 MHz, CD3CN) 7.50(d, J = 1.9 Hz, 1H), 7.41 (d, J = 1.6 Hz, 2H), 7.36 (dd, J = 1.7, 8.3Hz, 1H), 7.29-7.24 (m, 2H), 7.02 (dd, J = 2.1, 8.8 Hz, 1H), 6.24 (s,1H), 4.40 (t, J = 7.1 Hz, 2H), 3.80 (t, J = 7.1 Hz, 2H), 1.59- 1.55 (m,2H), 1.50 (s, 9H), 1.15-1.12 (m, 2H) 20 375.5 3.71 21 496 206 22 421.141.53 23 363.3 3.62 24 378.5 2.66 25 417.5 3.53 26 454.3 3.18 27 596.22.58 28 379.3 2.92 29 481 1.69 30 504.2 1.95 31 517 1.92 32 403.5 3.5 HNMR (400 MHz, DMSO) 10.76 (s, 1H), 8.72 (s, 1H), 7.79 (d, J = 2.3 Hz,1H), 7.62 (dd, J = 2.4, 8.6 Hz, 1H), 7.55 (d, J = 1.5 Hz, 1H), 7.14 (d,J = 8.6 Hz, 1H), 7.05-7.01 (m, 2H), 6.03 (d, J = 1.6 Hz, 1H), 4.54 (t, J= 6.4 Hz, 2H), 2.79 (t, J = 6.4 Hz, 2H), 1.44 (m, 2H), 1.32 (s, 9H),1.03 (m, 2H) 33 321.3 2.98 34 450.2 2.02 35 395.1 3.59 36 509 2.01 37447.2 2.02 38 379.1 2.16 H NMR (400 MHz, DMSO) 10.78 (s, 1H), 8.39 (s,1H), 7.57 (d, J = 1.7 Hz, 1H), 7.17 (d, J = 8.6 Hz, 1H), 7.03-6.90 (m,4H), 6.12 (d, J = 1.5 Hz, 1H), 6.03 (s, 2H), 5.18 (s, 1H), 1.50 (s, 6H),1.41-1.38 (m, 2H), 1.05- 0.97 (m, 2H) 39 373.3 3.74 40 372.8 3.8 41397.3 3.41 H NMR (400 MHz, DMSO) 11.44 (s, 1H), 8.52 (s, 1H), 7.85 (d, J= 1.2 Hz, 2H), 7.71 (d, J = 1.7 Hz, 1H), 7.47-7.43 (m, 2H), 7.32- 7.26(m, 2H), 7.12 (dd, J = 2.0, 8.7 Hz, 1H), 7.04 (d, J = 1.6 Hz, 1H),6.97-6.90 (m, 2H), 6.84 (d, J = 1.3 Hz, 1H), 6.03 (s, 2H), 1.43-1.40 (m,2H), 1.07-1.03 (m, 2H) 42 505.3 2.23 H NMR (400 MHz, DMSO- d6) 8.33 (s,1H), 7.52 (s, 1H), 7.42-7.39 (m, 2H), 7.33- 7.25 (m, 2H), 6.14 (s, 1H),4.99 (s, 1H), 4.31-4.27 (m, 3H), 3.64 (t, J = 7.0 Hz, 2H), 3.20 (t, J =7.6 Hz, 2H), 1.91 (t, J = 7.6 Hz, 2H), 1.46 (m, 2H), 1.39 (s, 6H), 1.13(m, 2H) 43 505.4 1.97 44 407.7 1.76 H NMR (400 MHz, DMSO) 10.31 (s, 1H),8.34 (s, 1H). 7.53 (d, J = 1.8 Hz, 1H), 7.03 (d, J = 1.6 Hz, 1H),6.97-6.90 (m, 3H), 6.05- 6.03 (m, 3H), 4.72 (s, 2H), 1.40-1.38 (m, 2H),1.34 (s, 9H), 1.04- 1.00(m, 2H) 45 497.2 2.26 46 391.3 3.41 47 377.53.48 48 427.5 4.09 49 402.2 3.06 50 421.1 1.81 51 407.5 3.34 52 464.32.87 53 405.3 3.65 54 375 1.84 55 505.4 1.96 56 335.3 3.18 57 445.2 3.2758 491 1.88 59 478 1.98 60 413.3 3.95 61 402.5 3.71 62 393.3 1.98 63407.2 2.91 64 505.4 1.98 65 377.5 3.53 66 417.5 4.06 67 333.3 3.53 68397.3 3.86 69 506 1.67 70 501 2.1 71 335.3 3.22 72 487 1.93 73 417.53.88 74 395 1.95 75 548 1.64 76 418.3 2.9 77 377.3 3.87 78 363.3 3.48 79476 1.8 80 447.3 2.18 81 492.4 2 82 564.3 1.35 83 467.3 1.72 84 445.23.08 85 389.5 3.86 86 374.3 3.11 87 435 3.87 88 465 1.89 89 411.3 3.8990 449.3 3.92 91 393.3 3.12 92 469.6 1.75 93 476.5 2.88 94 377.5 3.41 95375.3 3.43 H NMR (400 MHz, DMSO) 10.52 (s, 1H), 8.39 (s, 1H), 7.46 (d, J= 1.8 Hz, 1H), 7.10- 6.89 (m, 5H), 6.03 (s, 2H), 2.68-2.65 (m, 2H),2.56-2.54 (m, 2H), 1.82- 1.77 (m, 4H), 1.41-1.34 (m, 2H), 1.04-0.97 (m,2H) 96 346.1 3.1 97 367.3 3.72 98 440.3 3.26 99 393.1 3.18 H NMR (400MHz, DMSO- d6) 11.80 (s, 1H), 8.64 (s, 1H), 7.83 (m, 1H), 7.33-7.26 (m,2H), 7.07 (m, 1H), 7.02 (m, 1H), 6.96- 6.89 (m, 2H), 6.02 (s, 2H), 4.33(q, J = 7.1 Hz, 2H), 1.42- 1.39 (m, 2H), 1.33 (t, J = 7.1 Hz, 3H), 1.06-1.03 (m, 2H) 100 421.3 1.85 H NMR (400 MHz, DMSO) 13.05 (s, 1H), 9.96(d, J = 1.6 Hz, 1H), 7.89 (d, J = 1.9 Hz, 1H), 7.74 (d, J = 2.0 Hz, 1H),7.02 (d, J = 1.6 Hz, 1H), 6.96- 6.88 (m, 2H), 6.22 (d, J = 2.3 Hz, 1H),6.02 (s, 2H), 1.43- 1.40 (m, 2H), 1.37 (s, 9H), 1.06-1.02 (m, 2H) 101387.5 2.51 102 479 3.95 103 420.3 3.12 104 469.5 3.97 105 391.3 2.04 106375.2 2.82 107 349.3 3.33 108 503.3 1.88 109 451.5 1.59 110 361.5 3.7111 391.3 3.65 112 335.3 3.03 113 496.5 1.68 114 381.5 3.72 115 390.33.22 116 397.3 3.52 H NMR (400 MHz, DMSO- d6) 11.27 (d, J = 1.9 Hz, 1H),8.66 (s, 1H), 8.08 (d, J = 1.6 Hz, 1H), 7.65- 7.61 (m, 3H), 7.46-7.40(m, 2H), 7.31 (d, J = 8.7 Hz, 1H), 7.25-7.17 (m, 2H), 7.03 (d, J = 1.6Hz, 1H), 6.98-6.87 (m, 2H), 6.02 (s, 2H), 1.43-1.39 (m, 2H), 1.06- 1.02(m, 2H) 117 377.5 3.77 118 515.3 2.3 119 381.3 3.8 120 464.2 2.1 121 4651.74 122 395.2 3.74 123 383.3 3.52 124 388.5 3.56 125 411.3 3.85 126459.2 1.53 H NMR (400 MHz, CD3CN) 9.23 (s, 1H), 7.51-7.48 (m, 2H), 7.19(d, J = 8.6 Hz, 1H), 7.06-7.03 (m, 2H), 6.95-6.89 (m, 2H), 6.17 (dd, J =0.7, 2.2 Hz, 1H), 6.02 (s, 2H), 2.61- 2.57 (m, 2H), 2.07-2.03 (m, 2H),1.55-1.51 (m, 2H), 1.39 (s, 6H), 1.12- 1.09 (m, 2H) 127 408.5 2.48 128393 3.26 129 420.2 2.16 130 406.3 2.88 131 473.3 4.22 132 417.3 3.8 133465 1.74 134 464.3 2.91 135 347.3 3.42 136 511 2.35 137 455.5 3.29 138393.3 3.54 139 335.1 3.08 140 434.5 2.74 141 381.3 2.91 142 431.5 3.97143 539 1.89 144 515 1.89 145 407.5 3.6 146 379.5 1.51 147 409.3 4 148392.2 1.22 149 375.3 3.37 150 377.3 3.61 151 377.22 3.96 152 504.5 1.99153 393.1 3.47 154 363.3 3.52 155 321.3 3.13 156 407.5 3.2 157 406.31.43 158 379.3 1.89 159 451 3.34 160 375.3 3.82 161 355.1 3.32 162 4752.06 163 437.2 2.35 164 379.2 2.76 165 462 3.44 166 465.2 2.15 167 455.22.45 168 451 1.65 169 528 1.71 170 374.3 3.4 171 449.5 1.95 172 381.33.8 173 346.3 2.93 174 483.1 2.25 175 411.2 3.85 176 431.5 4.02 177485.5 4.02 178 528.5 1.18 179 473 1.79 180 479 2.15 181 387.5 2.56 182365.3 3.13 183 493 2.3 184 461.3 2.4 H NMR (400 MHz, DMSO- d6) 10.89 (s,1H), 8.29 (s, 1H), 7.52 (s, 1H), 7.42-7.37 (m, 2H), 7.32 (dd, J = 8.3,1.4 Hz, 1H), 7.01 (d, J = 10.9 Hz, 1H), 6.05 (d, J = 1.7 Hz, 1H), 4.29(t, J = 5.0 Hz, 1H), 3.23 (m, 2H), 1.81 (t, J = 7.7 Hz, 2H), 1.46 (m,2H), 1.29 (s, 6H), 1.13 (m, 2H) 185 377.5 3.63 186 464 1.46 187 339.13.2 188 435.5 1.64 189 392.3 2.18 190 435.5 3.67 H NMR (400 MHz, DMSO)11.83 (s, 1H), 10.76 (s, 1H), 8.53 (s, 1H), 7.93 (d, J = 1.8 Hz, 1H),7.60 (dd, J = 2.3, 8.5 Hz, 1H), 7.53 (d, J = 1.4 Hz, 1H), 7.14 (d, J =8.6 Hz, 1H), 7.02-6.97 (m, 2H), 6.02 (d, J = 1.5 Hz, 1H), 3.71 (t, J =6.2 Hz, 2H), 3.37 (t, J = 6.2 Hz, 2H), 3.25 (s, 3H), 1.44 (m, 2H), 1.32(s, 9H), 1.08 (m, 2H) 191 421.3 3.32 192 404.4 0.95 193 451 1.71 194 4651.69 195 434.2 2.29 196 363.3 3.4 197 501 1.91 198 411.2 3.14 199 4391.89 200 434.4 1.53 201 462 3.22 202 351.3 2.59 203 495.2 2.71 204 4353.94 205 397.3 3.69 206 493 2.26 207 487 1.87 208 391.3 2.94 209 397.23.3 210 487.2 1.85 H NMR (400 MHz, CD3CN) 7.50 (d, J = 2.0 Hz, 1H), 7.41(d, J = 1.6 Hz, 2H), 7.37-7.32 (m, 2H), 7.25 (d, J = 8.3 Hz, 1H), 6.98(dd, J = 2.1, 8.8 Hz, 1H), 6.27 (d, J = 0.6 Hz, 1H), 4.40-4.28 (m, 2H),4.12-4.06 (m, 1H), 3.59- 3.51 (m, 2H), 1.59-1.50 (m, 2H), 1.47 (s, 9H),1.15-1.12 (m, 2H) 211 381.3 3.69 212 461 2.04 213 469 1.72 214 363.33.48 215 432.3 3.07 216 403.5 3.94 217 420.4 1.27 218 475 2.2 219 484.31.84 220 419.3 3.87 221 486.3 0.91 222 391.3 3.01 223 398.3 1.3 224349.2 2.54 225 375.5 3.74 226 377.5 3.47 H NMR (400 MHz, DMSO- d6) 10.76(s, 1H), 8.39 (s, 1H), 7.55 (s, 1H), 7.15-7.13 (m, 1H), 7.03- 6.89 (m,4H), 6.03 (m, 3H), 1.41-1.38 (m, 2H), 1.32 (s, 9H), 1.04-1.01 (m, 2H)227 393.3 2.03 228 398.3 1.24 229 487.2 1.78 230 361.1 3.47 231 435.52.12 232 321.3 2.91 233 413.3 3.77 234 393.3 1.58 235 465 1.92 236 361.33.18 237 421 1.8 238 405.5 3.79 239 544.3 1.4 240 405.3 3.9 241 462 1.74242 550 1.68 243 395.2 1.98 244 517.3 1.94 245 372.2 3.59 246 361.3 3.58247 490 1.95 248 407.3 1.52 H NMR (400 MHz, DMSO) 10.74 (d, J = 1.2 Hz,1H), 8.40 (s, 1H), 7.54 (d, J = 1.8 Hz, 1H), 7.15 (d, J = 8.6 Hz, 1H),7.03-6.90 (m, 4H), 6.03- 6.00 (m, 3H), 3.26-3.22 (m, 2H), 1.85-1.80 (m,2H), 1.41- 1.38 (m, 2H), 1.31 (s, 6H), 1.05-1.01 (m, 2H) 249 393.3 3.32250 406.2 2.08 251 511 2.39 252 379.3 3.3 253 383 3.46 254 401.2 3.26255 398.3 1.38 256 512.5 1.96 257 389.2 3.05 258 321.3 3.02 259 392.12.74 260 462 1.81 261 453 1.91 262 349.3 3.22 263 391.1 3.67 H NMR (400MHz, DMSO) 1.01-1.05 (dd, J = 4.0, 6.7 Hz, 2H), 1.41-1.39 (m, 11H), 3.81(s, 3H), 6.03 (s, 2H), 6.15 (s, 1H), 6.96-6.90 (m, 2H), 7.02 (d, J = 1.6Hz, 1H), 7.09 (dd, J = 2.0, 8.8 Hz, 1H), 7.25 (d, J = 8.8 Hz, 1H), 7.60(d, J = 1.9 Hz, 1H), 8.46 (s, 1H) 264 421.3 1.66 H NMR (400 MHz, CD3CN)8.78 (s, 1H), 7.40 (m, 1H), 7.33 (s, 1H), 7.08 (m, 1H), 6.95-6.87 (m,3H), 6.79 (m, 1H), 5.91 (s, 2H), 3.51 (dd, J = 5.9, 7.8 Hz, 2H),2.92-2.88 (m, 2H), 2.64 (t, J = 5.8 Hz, 1H), 1.50 (m, 2H), 1.41 (s, 9H),1.06 (m, 2H) 265 475 2.15 266 347.3 3.32 267 420.5 1.81 268 416.2 1.76269 485 2.06 270 395.3 3.89 271 492 1.59 272 405.5 3.96 273 547.2 1.65274 631.6 1.91 275 590.4 2.02 276 465.7 1.79 277 411.3 2.14 278 385.31.99 279 425.3 2.19 280 473.2 1.74 281 469.4 2.02 H NMR (400 MHz, DMSO)8.82 (s, 1H), 7.84 (d, J = 1.7 Hz, 1H), 7.55- 7.51 (m, 2H), 7.40-7.35(m, 2H), 7.29 (dd, J = 1.7, 8.3 Hz, 1H), 7.04 (s, 1H), 4.98 (t, J = 5.6Hz, 1H), 4.27 (t, J = 6.1 Hz, 2H), 3.67 (q, J = 6.0 Hz, 2H), 1.48 (dd, J= 4.0, 6.7 Hz, 2H), 1.13 (dd, J = 4.1, 6.8 Hz, 2H) 282 644.4 1.83 283544.6 1.97 284 465.4 1.56 285 485.2 1.8 286 475.2 1.87 287 564.2 1.95288 512.5 1.89 H NMR (400 MHz, DMSO) 8.77 (s, 1H), 7.97 (s, 1H), 7.51(s, 1H), 7.43-7.40 (m, 2H), 7.33 (d, J = 8.2 Hz, 1H), 6.36 (s, 1H),4.99-4.97 (m, 2H), 4.52 (d, J = 13.1 Hz, 1H), 4.21 (dd, J = 9.2, 15.2Hz, 1H), 3.86 (m, 1H), 3.51-3.36 (m, 2H), 1.51- 1.48 (m, 2H), 1.43 (s,9H), 1.17-1.15 (m, 2H) 289 437.3 1.6 290 499.5 1.81 H NMR (400 MHz,DMSO) 8.82 (s, 1H), 7.83 (d, J = 1.7 Hz, 1H), 7.55- 7.50 (m, 2H),7.39-7.28 (m, 3H), 7.03 (s, 1H), 4.97 (d, J = 5.6 Hz, 1H), 4.83 (t, J =5.6 Hz, 1H), 4.33 (dd, J = 3.4, 15.1 Hz, 1H), 4.09 (dd, J = 8.7, 15.1Hz, 1H), 3.80-3.78 (m, 1H), 3.43- 3.38 (m, 1H), 3.35-3.30 (m, 1H),1.49-1.46 (m, 2H), 1.14- 1.11 (m, 2H) 291 455.4 2.02 H NMR (400 MHz,DMSO) 8.62 (s, 1H), 7.56 (s, 1H), 7.50 (s, 1H), 7.38 (d, J = 8.3 Hz,1H), 7.29 (dd, J = 1.5, 8.3 Hz, 1H), 7.23 (d, J = 8.7 Hz, 1H), 7.06 (dd,J = 1.7, 8.7 Hz, 1H), 6.19 (s, 1H), 4.86 (t, J = 5.4 Hz, 1H), 4.03 (t, J= 6.1 Hz, 2H), 3.73 (qn, J = 8.5 Hz, 1H), 3.57 (q, J = 5.9 Hz, 2H),2.39-2.33 (m, 2H), 2.18-1.98 (m, 3H), 1.88- 1.81 (m, 1H), 1.47-1.44 (m,2H), 1.11-1.09 (m, 2H) 292 578.4 1.99 293 630.4 1.8 294 443.4 1.98 H NMR(400 MHz, DMSO) 8.62 (s, 1H), 7.55 (d, J = 1.8 Hz, 1H), 7.50 (d, J = 1.5Hz, 1H), 7.38 (d, J = 8.3 Hz, 1H), 7.30-7.24 (m, 2H), 7.05 (dd, J = 2.0,8.8 Hz, 1H), 6.13 (s, 1H), 4.88 (t, J = 5.5 Hz, 1H), 4.14 (t, J = 6.1Hz, 2H), 3.61 (m, 2H), 3.21 (septet, J = 6.8 Hz, 1H), 1.47- 1.44 (m,2H), 1.26 (d, J = 6.8 Hz, 6H), 1.11- 1.08 (m, 2H) 295 482.3 2 H NMR (400MHz, DMSO) 8.78 (s, 1H), 7.92 (s, 1H), 7.51 (s, 1H), 7.45 (s, 1H), 7.41(d, J = 8.3 Hz, 1H), 7.33 (d, J = 8.4 Hz, 1H), 6.34 (s, 1H), 5.01 (t, J= 5.7 Hz, 1H), 4.41 (t, J = 6.6 Hz, 2H), 3.68 (m, 2H), 1.51- 1.47 (m,2H), 1.42 (s, 9H), 1.19-1.15 (m, 2H) 296 438.7 2.12 H NMR (400 MHz,DMSO) 11.43 (s, 1H), 8.74 (s, 1H), 7.63 (s, 1H), 7.51 (s, 1H), 7.45-7.40(m, 2H), 7.33 (dd, J = 1.4, 8.3 Hz, 1H), 6.25 (d, J = 1.5 Hz, 1H),1.51-1.48 (m, 2H), 1.34 (s, 9H), 1.17-1.14 (m, 2H) 297 449.3 1.6 298517.5 1.64 299 391.5 2.05 300 449.3 1.59 301 501.2 1.93 302 503.5 1.63303 437.3 1.6 304 425.1 2.04 H NMR (400 MHz, DMSO) 12.16 (s, 1H), 8.80(s, 1H), 7.83 (s, 1H), 7.51 (d, J = 1.4 Hz, 1H), 7.39- 7.28 (m, 4H),6.95 (s, 1H), 1.48 (dd, J = 4.0, 6.6 Hz, 2H), 1.13 (dd, J = 4.0, 6.7 Hz,2H) 305 459.2 1.67 306 558.4 2.05

VII. Assays for Detecting and Measuring ΔF508-CFTR Correction Propertiesof Compounds

Membrane Potential Optical Methods for Assaying ΔF508-CFTR ModulationProperties of Compounds

The optical membrane potential assay utilized voltage-sensitive FRETsensors described by Gonzalez and Tsien (See, Gonzalez, J. E. and R. Y.Tsien (1995) “Voltage sensing by fluorescence resonance energy transferin single cells” Biophys J 69(4): 1272-80, and Gonzalez, J. E. and R. Y.Tsien (1997) “Improved indicators of cell membrane potential that usefluorescence resonance energy transfer” Chem Biol 4(4): 269-77) incombination with instrumentation for measuring fluorescence changes suchas the Voltage/Ion Probe Reader (VIPR) (See, Gonzalez, J. E., K. Oades,et al. (1999) “Cell-based assays and instrumentation for screeningion-channel targets” Drug Discov Today 4(9): 431-439).

These voltage sensitive assays are based on the change in fluorescenceresonant energy transfer (FRET) between the membrane-soluble,voltage-sensitive dye, DiSBAC₂(3), and a fluorescent phospholipid,CC2-DMPE, which is attached to the outer leaflet of the plasma membraneand acts as a FRET donor. Changes in membrane potential (V_(m)) causethe negatively charged DiSBAC₂(3) to redistribute across the plasmamembrane and the amount of energy transfer from CC2-DMPE changesaccordingly. The changes in fluorescence emission were monitored usingVIPR™ II, which is an integrated liquid handler and fluorescent detectordesigned to conduct cell-based screens in 96- or 384-well microtiterplates.

Identification of Correction Compounds

To identify small molecules that correct the trafficking defectassociated with ΔF508-CFTR; a single-addition HTS assay format wasdeveloped. The cells were incubated in serum-free medium for 16 hrs at37° C. in the presence or absence (negative control) of test compound.As a positive control, cells plated in 384-well plates were incubatedfor 16 hrs at 27° C. to “temperature-correct” ΔF508-CFTR. The cells weresubsequently rinsed 3× with Krebs Ringers solution and loaded with thevoltage-sensitive dyes. To activate ΔF508-CFTR, 10 μM forskolin and theCFTR potentiator, genistein (20 μM), were added along with Cl⁻-freemedium to each well. The addition of Cl⁻-free medium promoted Cl⁻ effluxin response to ΔF508-CFTR activation and the resulting membranedepolarization was optically monitored using the FRET-basedvoltage-sensor dyes.

Identification of Potentiator Compounds

To identify potentiators of ΔF508-CFTR, a double-addition HTS assayformat was developed. During the first addition, a Cl⁻-free medium withor without test compound was added to each well. After 22 sec, a secondaddition of Cl⁻-free medium containing 2-10 μM forskolin was added toactivate ΔF508-CFTR. The extracellular Cl⁻ concentration following bothadditions was 28 mM, which promoted Cl− efflux in response to ΔF508-CFTRactivation and the resulting membrane depolarization was opticallymonitored using the FRET-based voltage-sensor dyes. Solutions BathSolution #1: (in mM) NaCl 160, KCl 4.5, CaCl₂ 2, MgCl₂ 1, HEPES 10, pH7.4 with NaOH.

Chloride-free bath solution: Chloride salts in Bath Solution #1 aresubstituted with gluconate salts.

CC2-DMPE: Prepared as a 10 mM stock solution in DMSO and stored at −20°C.

DiSBAC₂(3): Prepared as a 10 mM stock in DMSO and stored at −20° C.

Cell Culture

NIH3T3 mouse fibroblasts stably expressing ΔF508-CFTR are used foroptical measurements of membrane potential. The cells are maintained at37° C. in 5% CO₂ and 90% humidity in Dulbecco's modified Eagle's mediumsupplemented with 2 mM glutamine, 10% fetal bovine serum, 1×NEAA, β-ME,1× pen/strep, and 25 mM HEPES in 175 cm² culture flasks. For all opticalassays, the cells were seeded at 30,000/well in 384-well matrigel-coatedplates and cultured for 2 hrs at 37° C. before culturing at 27° C. for24 hrs. for the potentiator assay. For the correction assays, the cellsare cultured at 27° C. or 37° C. with and without compounds for 16-24hours Electrophysiological Assays for assaying ΔF508-CFTR modulationproperties of compounds Ussing Chamber Assay Ussing chamber experimentswere performed on polarized epithelial cells expressing ΔF508-CFTR tofurther characterize the ΔF508-CFTR modulators identified in the opticalassays. FRT^(ΔF508-CFTR) epithelial cells grown on Costar Snapwell cellculture inserts were mounted in an Ussing chamber (PhysiologicInstruments, Inc., San Diego, Calif.), and the monolayers werecontinuously short-circuited using a Voltage-clamp System (Department ofBioengineering, University of Iowa, Iowa, and, Physiologic Instruments,Inc., San Diego, Calif.). Transepithelial resistance was measured byapplying a 2-mV pulse. Under these conditions, the FRT epitheliademonstrated resistances of 4 KΩ/cm² or more. The solutions weremaintained at 27° C. and bubbled with air. The electrode offsetpotential and fluid resistance were corrected using a cell-free insert.Under these conditions, the current reflects the flow of Cl throughΔF508-CFTR expressed in the apical membrane. The I_(SC) was digitallyacquired using an MP100A-CE interface and AcqKnowledge software (v3.2.6;BIOPAC Systems, Santa Barbara, Calif.).

Identification of Correction Compounds

Typical protocol utilized a basolateral to apical membrane Cl⁻concentration gradient. To set up this gradient, normal ringer was usedon the basolateral membrane, whereas apical NaCl was replaced byequimolar sodium gluconate (titrated to pH 7.4 with NaOH) to give alarge Cl⁻ concentration gradient across the epithelium. All experimentswere performed with intact monolayers. To fully activate ΔF508-CFTR,forskolin (10 μM) and the PDE inhibitor, IBMX (100 μM), were appliedfollowed by the addition of the CFTR potentiator, genistein (50 μM).

As observed in other cell types, incubation at low temperatures of FRTcells stably expressing ΔF508-CFTR increases the functional density ofCFTR in the plasma membrane. To determine the activity of correctioncompounds, the cells were incubated with 10 μM of the test compound for24 hours at 37° C. and were subsequently washed 3× prior to recording.The cAMP- and genistein-mediated I_(SC) in compound-treated cells wasnormalized to the 27° C. and 37° C. controls and expressed as percentageactivity. Preincubation of the cells with the correction compoundsignificantly increased the cAMP- and genistein-mediated I_(SC) comparedto the 37° C. controls.

Identification of Potentiator Compounds

Typical protocol utilized a basolateral to apical membrane Cl⁻concentration gradient. To set up this gradient, normal ringers was usedon the basolateral membrane and was permeabilized with nystatin (360μg/ml), whereas apical NaCl was replaced by equimolar sodium gluconate(titrated to pH 7.4 with NaOH) to give a large Cl⁻ concentrationgradient across the epithelium. All experiments were performed 30 minafter nystatin permeabilization. Forskolin (10 μM) and all testcompounds were added to both sides of the cell culture inserts. Theefficacy of the putative ΔF508-CFTR potentiators was compared to that ofthe known potentiator, genistein.

Solutions

Basolateral solution (in mM): NaCl (135), CaCl₂ (1.2), MgCl₂ (1.2),K₂HPO₄ (2.4), KHPO₄ (0.6),N-2-hydroxyethylpiperazine-N′-2-ethanesulfonic acid (HEPES) (10), anddextrose (10). The solution was titrated to pH 7.4 with NaOH.

Apical solution (in mM): Same as basolateral solution with NaCl replacedwith Na Gluconate (135).

Cell Culture

Fisher rat epithelial (FRT) cells expressing ΔF508-CFTR(FRT^(ΔF508-CFTR)) were used for Ussing chamber experiments for theputative ΔF508-CFTR modulators identified from our optical assays. Thecells were cultured on Costar Snapwell cell culture inserts and culturedfor five days at 37° C. and 5% CO₂ in Coon's modified Ham's F-12 mediumsupplemented with 5% fetal calf serum, 100 U/ml penicillin, and 100μg/ml streptomycin. Prior to use for characterizing the potentiatoractivity of compounds, the cells were incubated at 27° C. for 16-48 hrsto correct for the ΔF508-CFTR. To determine the activity of correctionscompounds, the cells were incubated at 27° C. or 37° C. with and withoutthe compounds for 24 hours.

The macroscopic ΔF508-CFTR current (I_(ΔF508)) in temperature- and testcompound-corrected NIH3T3 cells stably expressing ΔF508-CFTR weremonitored using the perforated-patch, whole-cell recording. Briefly,voltage-clamp recordings of I_(ΔF508) were performed at room temperatureusing an Axopatch 200B patch-clamp amplifier (Axon Instruments Inc.,Foster City, Calif.). All recordings were acquired at a samplingfrequency of 10 kHz and low-pass filtered at 1 kHz. Pipettes had aresistance of 5-6 MΩ when filled with the intracellular solution. Underthese recording conditions, the calculated reversal potential for Cl⁻(E_(Cl)) at room temperature was −28 mV. All recordings had a sealresistance >20 GΩ and a series resistance <15 M. Pulse generation, dataacquisition, and analysis were performed using a PC equipped with aDigidata 1320 A/D interface in conjunction with Clampex 8 (AxonInstruments Inc.). The bath contained <250 μl of saline and wascontinuously perifused at a rate of 2 ml/min using a gravity-drivenperfusion system.

Identification of Correction Compounds

To determine the activity of correction compounds for increasing thedensity of functional ΔF508-CFTR in the plasma membrane, we used theabove-described perforated-patch-recording techniques to measure thecurrent density following 24-hr treatment with the correction compounds.To fully activate ΔF508-CFTR, 10 μM forskolin and 20 μM genistein wereadded to the cells. Under our recording conditions, the current densityfollowing 24-hr incubation at 27° C. was higher than that observedfollowing 24-hr incubation at 37° C. These results are consistent withthe known effects of low-temperature incubation on the density ofΔF508-CFTR in the plasma membrane. To determine the effects ofcorrection compounds on CFTR current density, the cells were incubatedwith 10 μM of the test compound for 24 hours at 37° C. and the currentdensity was compared to the 27° C. and 37° C. controls (% activity).Prior to recording, the cells were washed 3× with extracellularrecording medium to remove any remaining test compound. Preincubationwith 10 μM of correction compounds significantly increased the cAMP- andgenistein-dependent current compared to the 37° C. controls.

Identification of Potentiator Compounds

The ability of ΔF508-CFTR potentiators to increase the macroscopicΔF508-CFTR Cl current (I_(ΔF508)) in NIH3T3 cells stably expressingΔF508-CFTR was also investigated using perforated-patch-recordingtechniques. The potentiators identified from the optical assays evoked adose-dependent increase in I_(ΔF508) with similar potency and efficacyobserved in the optical assays. In all cells examined, the reversalpotential before and during potentiator application was around −30 mV,which is the calculated E_(Cl) (˜28 mV).

Solutions

Intracellular solution (in mM): Cs-aspartate (90), CsCl (50), MgCl₂ (1),HEPES (10), and 240 μg/ml amphotericin-B (pH adjusted to 7.35 withCsOH).

Extracellular solution (in mM): N-methyl-D-glucamine (NMDG)-Cl (150),MgCl₂ (2), CaCl₂ (2), HEPES (10) (pH adjusted to 7.35 with HCl).

Cell Culture

NIH3T3 mouse fibroblasts stably expressing ΔF508-CFTR are used forwhole-cell recordings. The cells are maintained at 37° C. in 5% CO₂ and90% humidity in Dulbecco's modified Eagle's medium supplemented with 2mM glutamine, 10% fetal bovine serum, 1× NEAA, β-ME, 1× pen/strep, and25 mM HEPES in 175 cm² culture flasks. For whole-cell recordings,2,500-5,000 cells were seeded on poly-L-lysine-coated glass coverslipsand cultured for 24-48 hrs at 27° C. before use to test the activity ofpotentiators; and incubated with or without the correction compound at37° C. for measuring the activity of correctors.

The single-channel activities of temperature-corrected ΔF508-CFTR stablyexpressed in NIH3T3 cells and activities of potentiator compounds wereobserved using excised inside-out membrane patch. Briefly, voltage-clamprecordings of single-channel activity were performed at room temperaturewith an Axopatch 200B patch-clamp amplifier (Axon Instruments Inc.). Allrecordings were acquired at a sampling frequency of 10 kHz and low-passfiltered at 400 Hz. Patch pipettes were fabricated from Corning KovarSealing #7052 glass (World Precision Instruments, Inc., Sarasota, Fla.)and had a resistance of 5-8 MΩ when filled with the extracellularsolution. The ΔF508-CFTR was activated after excision, by adding 1 mMMg-ATP, and 75 nM of the cAMP-dependent protein kinase, catalyticsubunit (PKA; Promega Corp. Madison, Wis.). After channel activitystabilized, the patch was perifused using a gravity-drivenmicroperfusion system. The inflow was placed adjacent to the patch,resulting in complete solution exchange within 1-2 sec. To maintainΔF508-CFTR activity during the rapid perifusion, the nonspecificphosphatase inhibitor F (10 mM NaF) was added to the bath solution.Under these recording conditions, channel activity remained constantthroughout the duration of the patch recording (up to 60 min). Currentsproduced by positive charge moving from the intra- to extracellularsolutions (anions moving in the opposite direction) are shown aspositive currents. The pipette potential (V_(p)) was maintained at 80mV.

Channel activity was analyzed from membrane patches containing ≤2 activechannels. The maximum number of simultaneous openings determined thenumber of active channels during the course of an experiment. Todetermine the single-channel current amplitude, the data recorded from120 sec of ΔF508-CFTR activity was filtered “off-line” at 100 Hz andthen used to construct all-point amplitude histograms that were fittedwith multigaussian functions using Bio-Patch Analysis software(Bio-Logic Comp. France). The total microscopic current and openprobability (P.) were determined from 120 sec of channel activity. TheP_(o) was determined using the Bio-Patch software or from therelationship P_(o)=I/i(N), where I=mean current, i=single-channelcurrent amplitude, and N=number of active channels in patch.

Solutions

Extracellular solution (in mM): NMDG (150), aspartic acid (150), CaCl₂(5), MgCl₂ (2), and HEPES (10) (pH adjusted to 7.35 with Tris base).

Intracellular solution (in mM): NMDG-Cl (150), MgCl₂ (2), EGTA (5), TES(10), and Tris base (14) (pH adjusted to 7.35 with HCl).

Cell Culture

NIH3T3 mouse fibroblasts stably expressing ΔF508-CFTR are used forexcised-membrane patch-clamp recordings. The cells are maintained at 37°C. in 5% CO₂ and 90% humidity in Dulbecco's modified Eagle's mediumsupplemented with 2 mM glutamine, 10% fetal bovine serum, 1×NEAA, β-ME,1× pen/strep, and 25 mM HEPES in 175 cm² culture flasks. For singlechannel recordings, 2,500-5,000 cells were seeded onpoly-L-lysine-coated glass coverslips and cultured for 24-48 hrs at 27°C. before use.

Compounds of the invention are useful as modulators of ATP bindingcassette transporters. Using the procedures described above, theactivities, i.e., EC50s, of compounds of the present invention have beenmeasured to be from about 3.8 nM to about 13.5 μM. Furthermore, usingthose methods described above, the efficacies of compounds of thepresent invention have been measured to be from about 35% to about 110%.

OTHER EMBODIMENTS

It is to be understood that while the invention has been described inconjunction with the detailed description thereof, the foregoingdescription is intended to illustrate and not limit the scope of theinvention, which is defined by the scope of the appended claims. Otheraspects, advantages, and modifications are within the scope of thefollowing claims.

1-40. (canceled)
 41. A compound of formula IIc:

or a pharmaceutically acceptable salt thereof, wherein: R₁ is —Z^(A)R₄,wherein each Z^(A) is independently a bond or an optionally substitutedbranched or straight C₁₋₆ aliphatic chain, wherein up to two carbonunits of Z^(A) are optionally and independently replaced by —CO—, —CS—,—CONR^(A)—, —CONR^(A)NR^(A)—, —CO₂—, —OCO—, —NR^(A)CO₂, —O—,—NR^(A)CONR^(A)—, —OCONR^(A)—, —NR^(A)NR^(A)—, —NR^(A)CO—, —S—, —SO—,—SO₂—, —NR^(A)—, —SO₂NR^(A)—, —NR^(A)SO₂, or —NR^(A)SO₂NR^(A)—; and eachR₄ is independently R^(A), halo, —OH, —NH₂, —NO₂, —CN, or —OCF₃; andwherein each R^(A) is independently selected from hydrogen, anoptionally substituted aliphatic, an optionally substitutedcycloaliphatic, an optionally substituted heterocycloaliphatic, anoptionally substituted aryl, and an optionally substituted heteroaryl;each R₂ is —Z^(B)R₅, wherein each Z^(B) is independently a bond or anoptionally substituted branched or straight C₁₋₆ aliphatic chain,wherein up to two carbon units of Z^(B) are optionally and independentlyreplaced by —CO—, —CS—, —CONR^(B)—, —CONR^(B)NR^(B)—, —CO₂—, —OCO—,—NR^(B)CO₂, —O—, —NR^(B)CONR^(B)—, —OCONR^(B)—, —NR^(B)NR^(B)—,—NR^(B)CO—, —S—, —SO—, —SO₂—, —NR^(B)—, —SO₂NR^(B)—, —NR^(B)SO₂, or—NR^(B)SO₂NR^(B)—; and each R₅ is independently selected from R^(B),halogen, —OH, —NH₂, —NO₂, —CN, CF₃, and —OCF₃; and wherein each R^(B) isindependently selected from hydrogen, an optionally substitutedaliphatic, an optionally substituted cycloaliphatic, an optionallysubstituted heterocycloaliphatic, an optionally substituted aryl, and anoptionally substituted heteroaryl; or wherein any two adjacent R₂groups, together with the atoms to which they are attached, form anoptionally substituted carbocycle or an optionally substitutedheterocycle; and each R₃ and R′3 is independently —Z^(C)R₆; wherein eachZ^(C) is independently a bond or an optionally substituted branched orstraight C₁₋₆ aliphatic chain, wherein up to two carbon units of Z^(C)are optionally and independently replaced by —CO—, —CS—, —CONR^(C)—,—CONR^(C)NR^(C)—, —CO₂—, —OCO—, —NR^(C)CO₂, —O—, —NR^(C)CONR^(C)—,—OCONR^(C)—, —NR^(C)NR^(C)—, —NR^(C)CO—, —S—, —SO—, —SO₂—, —NR^(C)—,—SO₂NR^(C)—, —NR^(C)SO₂, or —NR^(C)SO₂NR^(C)—; each R₆ is independentlyselected from R^(C), halogen, —OH, —NH₂, —NO₂, —CN, and —OCF₃; andwherein each R^(C) is independently selected from hydrogen, anoptionally substituted aliphatic, an optionally substitutedcycloaliphatic, an optionally substituted heterocycloaliphatic, anoptionally substituted aryl, and an optionally substituted heteroaryl;or wherein any two adjacent R₃ groups, together with the atoms to whichthey are attached, form an optionally substituted carbocycle or anoptionally substituted heterocycle; n is 1-3; and p is 0-3.
 42. Acompound of formula IId:

or a pharmaceutically acceptable salt thereof; wherein: both R₂ groups,together with the atoms to which they are attached, form a groupselected from:

R′₃ is independently selected from: —H, —CH₃, —CH₂CH₃, —C(O)CH₃,—CH₂CH₂OH, —C(O)OCH₃,

 and each R₃ is independently selected from —H, —CH₃, —CH₂OH, —CH₂CH₃,—CH₂CH₂OH, —CH₂CH₂CH₃, —NH₂, halo, —OCH₃, —CN, —CF₃, —C(O)OCH₂CH₃,—S(O)₂OCH₃, —CH₂NH₂, —C(O)NH₂,


43. A compound according to claim 42, wherein both R₂ groups, togetherwith the atoms to which they are attached, is selected from the groups:


44. A compound according to claim 43, wherein R′₃ is hydrogen or thegroup:

wherein R₃₁ is H or a C₁₋₂ aliphatic that is optionally substituted with1-3 halogen, —OH, or combinations thereof, R₃₂ is -L-R₃₃; and wherein -Lis a bond, —CH₂—, —CH₂O—, —CH₂NHS(O)₂, —CH₂C(O)—, —CH₂NHC(O)—, or—CH₂NH—; and —R₃₃ is hydrogen, C₁₋₂ aliphatic, cycloaliphatic,heterocycloaliphatic, or heteroaryl, each of which is optionallysubstituted with one of —OH, —NH₂, or —CN.
 45. A compound according toclaim 44, wherein R₃₁ is hydrogen and R₃₂ is C₁₋₂ aliphatic optionallysubstituted with one of —OH, NH₂, or —CN.
 46. A pharmaceuticalcomposition comprising a compound or a pharmaceutically acceptable saltthereof according to any one of claims 41-44, and a pharmaceuticallyacceptable carrier, adjuvant, or vehicle.
 47. A pharmaceuticalcomposition according to claim 46, wherein the pharmaceuticalcomposition further comprises one or more additional therapeutic agents.48. A pharmaceutical composition comprising a compound or apharmaceutically acceptable salt thereof according to claim 45, and apharmaceutically acceptable carrier, adjuvant, or vehicle.
 49. Apharmaceutical composition according to claim 48, wherein thepharmaceutical composition further comprises one or more additionaltherapeutic agents.
 50. A method of treating or lessening the severityof cystic fibrosis in a patient, comprising the step of administering toa patient an effective amount of a compound or a pharmaceuticallyacceptable salt thereof according to any one of claims 41-44.
 51. Amethod of treating or lessening the severity of cystic fibrosis in apatient, comprising the step of administering to a patient an effectiveamount of a compound or a pharmaceutically acceptable salt thereofaccording to claim 45.